NMDA receptor

Mice with learning impairments caused by Aβ 25–35 exhibit anti-amnesic effects when given fluoroethylnormemantine (0.1–10 mg/kg; a single ip)[1]. In mice, Aβ 25-35-induced behavioral impairments, neuroinflammation, oxidative stress, apoptosis, and cell death are attenuated by fluoroethylnormemantine (0.1–10 mg/kg; ip once daily for 7 days)[1]. In rats, fluoroethylnormemantine (1–20 mg/kg; single injection) lowers fear behavior in cued fear conditioning (FC) and extinction training, as well as behavioral despair in the forced swim test (FST)[2].

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Given after stress, FluoroethylnormemantineFENM decreased behavioral despair and reduced perseverative behavior. When administered after re-exposure, FENM facilitated extinction learning. As a prophylactic, FENM attenuated learned fear and decreased stress-induced behavioral despair. FENM was behaviorally effective in both male and female mice. (R,S)-ketamine, but not FENM, increased expression of c-fos in vCA3. Both (R,S)-ketamine and FENM attenuated large-amplitude AMPA receptor–mediated bursts in vCA3, indicating a common neurobiological mechanism for further study. Conclusions Our results indicate that Fluoroethylnormemantine/FENM is a novel drug that is efficacious when administered at various times before or after stress. Future work will further characterize FENM’s mechanism of action with the goal of clinical development. [1]

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Results: Unlike memantine, Fluoroethylnormemantine/FENM did not produce nonspecific side effects and did not alter sensorimotor gating or locomotion. FENM decreased immobility in the forced swim test. Moreover, FENM robustly facilitated fear extinction learning when administered prior to either cued fear conditioning training or tone reexposure. Conclusions: These results suggest that FENM is a promising, novel compound that robustly reduces fear behavior and may be useful for further preclinical testing. [2]

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Results: Both Memantine and FluoroethylnormemantineFENM showed symptomatic anti-amnesic effects in Aβ 25-35-treated mice. Interestingly, FENM was not amnesic when tested alone at 10 mg/kg, contrarily to Memantine. Drugs injected once per day prevented Aβ 25-35-induced memory deficits, oxidative stress (lipid peroxidation, cytochrome c release), inflammation (interleukin-6, tumor necrosis factor-α increases; glial fibrillary acidic protein and Iba1 immunoreactivity in the hippocampus and cortex), and apoptosis and cell loss (Bcl-2-associated X/B-cell lymphoma 2 ratio; cell loss in the hippocampus CA1 area). However, FENM effects were more robust than observed with Memantine, with significant attenuations vs the Aβ 25-35-treated group. Conclusions: FENM therefore appeared as a potent neuroprotective drug in an AD model, with a superior efficacy compared with Memantine and an absence of direct amnesic effect at higher doses. These results open the possibility to use the compound at more relevant dosages than those actually proposed in Memantine treatment for AD [3].

Animal/Disease Models: Male Swiss CD-1 mice (7-9 weeks) were injected with Aβ25-35[1]
Doses: 0.1, 0.3, 1, 3, 10 mg/kg
Route of Administration: Ip 30 minutes before the behavioral tests
Experimental Results: Attenuated Aβ 25-35-induced spontaneous alternation deficit, passive avoidance deficit, and novel object exploration deficit.

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Fluoroethylnormemantine: FENM/Fluoroethylnormemantine was administered in a single dose at 10, 20, or 30 mg/kg of body weight. Saline, memantine (10 mg/kg), (R,S)-ketamine (30 mg/kg), or FENM (10, 20, or 30 mg/kg) was administered before or after contextual fear conditioning in 129S6/SvEv mice. Drug efficacy was assayed using various behavioral tests. Protein expression in the hippocampus was quantified with immunohistochemistry or Western blotting. In vitro radioligand binding was used to assay drug binding affinity. Patch clamp electrophysiology was used to determine the effect of drug administration on glutamatergic activity in ventral hippocampal cornu ammonis 3 (vCA3) 1 week after injection. [1]

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Researchers administered saline, FENM, or memantine prior to a number of behavioral assays, including paired-pulse inhibition, open field, light dark test, forced swim test, and cued fear conditioning in male Wistar rats. FENM/Fluoroethylnormemantine was administered in a single dose at 1, 3, 5, 10, or 20 mg/kg of body weight. [2]

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Swiss mice were treated intracerebroventricularly with aggregated Aβ 25-35 peptide and examined after 1 week in a battery of memory tests (spontaneous alternation, passive avoidance, object recognition, place learning in the water-maze, topographic memory in the Hamlet). Toxicity induced in the mouse hippocampus or cortex was analyzed biochemically or morphologically.
Researchers examined 2 effects of the drugs/Fluoroethylnormemantine. First, symptomatic effects were analyzed in Aβ 25-35-treated mice by injecting the drugs just before the behavioral tests. Second, the neuroprotection was analyzed by repeatedly o.d. injecting the mice for 1 week starting on the day of peptide injection. For symptomatic effects, drugs were injected only on day 8 after Aβ 25–35 injection, 30 minutes before the behavioral tests: spontaneous alternation, passive avoidance training, session 2 of the object recognition test or each water-maze training sessions (supplementary Figure 1a). A group was tested for spontaneous alternation, passive avoidance, and object recognition in series. As Memantine, and expectedly Fluoroethylnormemantine/FENM, has a short half-life in mice (<2 hours; Beconi et al., 2011), all the drug was excreted overnight. A separate group was trained in the Hamlet before Aβ 25–35 injection to assess topographic memory (supplementary Figure 1b). For neuroprotective effects, drugs were injected o.d. from day 1 to day 7 after Aβ 25–35 injection (supplementary Figure 1c), and mice were tested for spontaneous alternation, passive avoidance, and object recognition in series. They were killed at day 13 for immunochemistry (group A). A group of mice performed place learning in the water-maze, then were killed at day 16 and used for biochemical assays (group B). An additional series (group C) included mice killed at day 5 after Aβ 25–35 peptide injection and daily drug injections for assessing cytokine levels by enzyme-linked immuno-sorbent assays (ELISA).[3]

Recently, a novel NMDAR antagonist, fluoroethyl normemantine (FENM), was derived from the NMDAR antagonist memantine. To determine its biodistribution and safety, researchers developed FENM into a radiolabeled compound, [18F]-FENM. In rats, brain drug concentrations stabilized 40 minutes after injection of [18F]-FENM (44 ± 11 MBq), with the brain concentration representing 0.4% of the injected dose. In rats anesthetized with isoflurane prior to [18F]-FENM injection, in vitro autoradiography and immunohistochemical staining combined with other methods showed that NMDAR and [18F]-FENM binding sites were highly colocalized, particularly in the cortical and hippocampal (HPC) regions. Most interestingly, if rats were anesthetized immediately before [18F]-FENM injection with (R,S)-ketamine (80 mg/kg), the autoradiographic signal of [18F]-FENM was no longer correlated with NMDAR staining, indicating that its binding was inhibited or blocked. [18F]-FENM has a Ki value of 3.5 μM, while (R,S)-ketamine has a Ki value of 0.53 μM. In addition, recent studies have shown that FENM can promote extinction learning in male rats without affecting their sensorimotor behavior. However, whether FENM has preventive or antidepressant effects remains to be studied. [1] Recently, a novel radiolabeled compound, [18F]-fluoroethyl normethmannet (FENM), derived from memantine, has been used as a novel positron emission tomography (PET) tracer (Salabert et al., 2015, 2018). [18F]-FENM has a Ki value of 3.5 × 10⁻⁶ M, is highly lipophilic (logD = 1.93), stabilizes within 40 minutes after injection, and has a residual amount in the brain of about 0.4% of the initial dose. The results of ex vivo autoradiography and immunohistochemistry showed that [18F]-FENM was highly colocalized with NMDARs in the cortex and cerebellum. Interestingly, injection of (R,S)-ketamine blocked this co-localization, suggesting that FENM has a lower affinity for the NMDAR receptor than (R,S)-ketamine. Furthermore, since both compounds bind to the phencycline localization site in the NMDAR channel pores, these data suggest they may also have similar behavioral effects. However, although the antidepressant-like effects of FENM remain unclear, recent data indicate that FENM can enhance cognitive function and exert neuroprotective effects in an AD mouse model (Couly et al., 2020). In this study, Couly and colleagues found that FENM reversed deficits in long-term memory, navigation, spatial learning, and object recognition in a pharmacological model of AD. Interestingly, compared to memantine, the authors found that FENM improved spatiotemporal orientation in mice on the Hamlet test, while memantine did not affect the mice's behavior. The study found that the behavioral effects of FENM corresponded to a reduction in inflammatory cytokines and loss of neurons in the hippocampus. Therefore, although FENM has shown potential to enhance cognitive function and protect against age-related brain damage, it remains unclear whether the drug can reverse stress-related maladaptive behaviors. [2]

[1]. Anti-Amnesic and Neuroprotective Effects of Fluoroethylnormemantine in a Pharmacological Mouse Model of Alzheimer's Disease. Int J Neuropsychopharmacol. 2021 Feb 15;24(2):142-157.

[2]. Fluoroethylnormemantine, a novel derivative of memantine, facilitates extinction learning without sensorimotor deficits. Int J Neuropsychopharmacol. 2021 Jul 14;24(6):519-531.

[3]. Fluoroethylnormemantine, a novel NMDA receptor antagonist, for the prevention and treatment of stress-induced maladaptive behavior. Biological Psychiatry. 2021 Feb 15;24(2):142-157.

This study characterized FENM, an NMDAR antagonist with antidepressant and preventive effects. We found that: 1) FENM exhibited antidepressant-like properties in both male and female mice under stress; 2) FENM inhibited loss of appetite in non-stressed male mice; 3) FENM administration after extinction in male mice reduced fear; 4) Administration of FENM one week before stress exposure prevented stress in both male and female mice; 5) One week after FENM administration reduced burst discharge mediated by large-amplitude AMPA receptors in the vCA3 region of the hippocampus. [1] This study aimed to investigate the effects of FENM administration on sensorimotor gating, avoidance behavior, behavioral despair, and fear behavior in rats. The results showed that FENM effectively reduced behavioral despair and promoted extinction learning without affecting motor or sensorimotor behavior. These results indicate that FENM has antidepressant-like effects and a fear-reducing effect. FENM also effectively reduced learned fear when administered acutely before fear conditioning (FC), suggesting that it may also be used as a preventative drug to enhance resilience. In summary, our results suggest that FENM, as a novel NMDAR antagonist, may be suitable for further preclinical trials in a variety of stress-related behavioral tests. [2]

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FENM appears to be a promising drug. Its efficacy must now be validated in transgenic mouse models of Alzheimer's disease (AD). Only through repeated administration in these chronic models can it be determined whether FENM can reduce amyloid load and plaque formation in amyloid models, or reduce kinase activity and neurofibrillary tangle formation in tau models. Memantine (Wang et al., 2015) and several other drugs with similar symptom-improving and neuroprotective effects have been reported previously. In the future, the strength of the neuroprotective effect induced by FENM needs to be studied in similar transgenic models, and its mechanism of action needs to be analyzed to determine whether the molecule is superior to memantine and whether the drug can be used as a potential candidate for synergistic combination with drugs currently under development. In summary, we describe the symptom relief and neuroprotective efficacy of a novel memantine derivative, FENM, in a mouse model of Alzheimer's disease (AD). Compared with the parent molecule, FENM is more effective in preventing oxidative stress, apoptosis and neuroinflammation, suggesting that the molecule can not only be used as an effective PET radiotracer for NMDAR, but also has the potential to become a neuroprotective drug for AD. In addition, the compound may be used at a more appropriate dose than the dose currently recommended for memantine treatment. [3]

C12H20FN

197.29

197.157

C, 73.05; H, 10.22; F, 9.63; N, 7.10

1639210-26-6

Fluoroethylnormemantine hydrochloride;1639210-25-5

170907856

Colorless to light yellow ointment

3.2

1

2

2

14

237

2

C1[C@@H]2CC3(C[C@H]1CC(C2)(C3)N)CCF

HUYVZSFADWYSHD-ZYANWLCNSA-N

InChI=1S/C12H20FN/c13-2-1-11-4-9-3-10(5-11)7-12(14,6-9)8-11/h9-10H,1-8,14H2/t9-,10+,11?,12?

(5S,7R)-3-(2-fluoroethyl)adamantan-1-amine

Fluoroethylnormemantine; SCHEMBL16274382;

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.)