Enzymes of porcine brain hydrolyzing 2-arachidonoylglycerol, an endogenous ligand of cannabinoid receptors

Goparaju, SravanKumar; Ueda, Natsuo; Taniguchi, Kyoko; Yamamoto, Shozo

doi:10.1016/s0006-2952(98)00314-1

Cited by 207 publications

(163 citation statements)

References 41 publications

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“…As further evident from Figure 5a, MAFP was significantly more effective than PMSF, although it was used at a 100-fold smaller concentration. This is consistent with previous findings demonstrating that MAFP potently (IC 50 B3 nM) inhibits brain 2-AG hydrolzing enzymatic activity (Goparaju et al, 1999). Indeed, our recent work indicates that MAFP is B70,000-fold more potent than PMSF in inhibiting 2-AG hydrolyzing activity in this preparate (S.M.…”

Section: Jr Savinainen Et Alsupporting

confidence: 93%

“…Optimized method for endocannabinoid activity 1457 (Goparaju et al, 1999). Contrary to this status, MAFP has also been described as an irreversible CB 1 receptor antagonist (Fernando & Pertwee, 1997).…”

Section: Jr Savinainen Et Almentioning

confidence: 99%

“…The degradation of 2-AG was substantially inhibited by pretreatment of membranes with the nonspecific serine protease inhibitor phenylmethylsulfonyl fluoride (PMSF) (Savinainen et al, 2001). Other studies have revealed that methyl arachidonoyl fluorophosphonate (MAFP) is clearly a more potent inhibitor of MGL and FAAH, than PMSF (Deutsch et al, 1997;Goparaju et al, 1999). In spite of this, MAFP is not being commonly used as an inhibitor of endocannabinoid degradation in G-protein-activation studies.…”

Section: Introductionmentioning

confidence: 99%

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An optimized approach to study endocannabinoid signaling: evidence against constitutive activity of rat brain adenosine A₁ and cannabinoid CB₁ receptors

Savinainen¹,

Saario²,

Niemi³

et al. 2003

British J Pharmacology

106

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1 At nanomolar concentrations, SR141716 and AM251 act as specific and selective antagonists of the cannabinoid CB 1 receptor. In the micromolar range, these compounds were shown to inhibit basal G-protein activity, and this is often interpreted to implicate constitutive activity of the CB 1 receptors in native tissue. We show here, using [ 35 S]GTPgS binding techniques, that micromolar concentrations of SR141716 and AM251 inhibit basal G-protein activity in rat cerebellar membranes, but only in conditions where tonic adenosine A 1 receptor signaling is not eliminated. 2 Unlike lipophilic A 1 receptor antagonists (potency order DPCPXbN-0840 Ecirsimarin4caf-feine), adenosine deaminase (ADA) was not fully capable in eliminating basal A 1 receptor-dependent G-protein activity. Importantly, all antagonists reduced basal signal to the same extent (20%), and the response evoked by the inverse agonist DPCPX was not reversed by the neutral antagonist N-0840. These data indicate that rat brain A 1 receptors are not constitutively active, but that an ADA-resistant adenosine pool is responsible for tonic A 1 receptor activity in brain membranes. 3 SR141716 and AM251, at concentrations fully effective in reversing CB 1 -mediated responses (10 À6 M), did not reduce basal G-protein activity, indicating that CB 1 receptors are not constitutively active in these preparations. 4 At higher concentrations (1 -2.5 Â 10 À5 M), both antagonists reduced basal G-protein activity in control and ADA-treated membranes, but had no effect when A 1 receptor signaling was blocked with DPCPX. Moreover, the CB 1 antagonists right-shifted A 1 agonist dose -response curves without affecting maximal responses, suggesting competitive mode of antagonist action. The CB 1 antagonists did not affect muscarinic acetylcholine or GABA B receptor signaling. 5 When further optimizing G-protein activation assay for the labile endocannabinoid 2-arachidonoylglycerol (2-AG), we show, by using HPLC, that pretreatment of cerebellar membranes with methyl arachidonoyl fluorophosphonate (MAFP) fully prevented enzymatic degradation of 2-AG and concomitantly enhanced the potency of 2-AG. In contrast to previous claims, MAFP exhibited no antagonist activity at the CB 1 receptor. 6 The findings establish an optimized method with improved signal-to-noise ratio to assess endocannabinoid-dependent G-protein activity in brain membranes, under assay conditions where basal adenosinergic tone and enzymatic degradation of 2-AG are fully eliminated.

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Section: Jr Savinainen Et Alsupporting

confidence: 93%

Section: Jr Savinainen Et Almentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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An optimized approach to study endocannabinoid signaling: evidence against constitutive activity of rat brain adenosine A₁ and cannabinoid CB₁ receptors

Savinainen¹,

Saario²,

Niemi³

et al. 2003

British J Pharmacology

106

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“…The key enzymes responsible for the hydrolysis of endocannabinoids are fatty acid amide hydrolase (FAAH) and monoglyceride lipase (MGL, EC 3.1.1.23) [13,14]. FAAH is mainly responsible for hydrolysis of AEA to arachidonic acid and ethanolamine, and MGL for hydrolysis of 2-AG to arachidonic acid and glycerol [15][16][17]. Numerous potent inhibitors against FAAH have been reported, including compounds that have shown promising in vivo activity, selectivity and therapeutic effects [18][19][20][21].…”

Section: Introductionmentioning

confidence: 99%

Chiral 3-(4,5-dihydrooxazol-2-yl)phenyl alkylcarbamates as novel FAAH inhibitors: Insight into FAAH enantioselectivity by molecular docking and interaction fields

Myllymäki¹,

Käsnänen

Kataja³

et al. 2009

European Journal of Medicinal Chemistry

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Fatty acid amide hydrolase (FAAH) and monoglyceride lipase (MGL) are the main enzymes responsible for the hydrolysis of endogenous cannabinoids N-arachidonoylethanolamide (AEA) and 2-arachidonoylglycerol (2-AG), respectively. Phenyl alkylcarbamates are FAAH inhibitors with anxiolytic and analgesic activities in vivo. Herein we present for the first time the synthesis and biological evaluation of a series of chiral 3-(2-oxazoline)-phenyl N-alkylcarbamates as FAAH inhibitors. Furthermore, the structural background of chirality on the FAAH inhibition is explored by analyzing the protein-ligand interactions. Remarkably, 10-fold difference in potency was observed for (R)-and (S)-derivatives of 3-(5-methyl-4,5-dihydrooxazol-2-yl)phenyl cyclohexylcarbamate (6a vs. 6b). Molecular modelling indicated an important interaction between the oxazoline nitrogen and FAAH active site.

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“…This appears to be unlikely, however, for three reasons. First, pig brain tissue contains two distinct 2-AG-hydrolase activities, both of which are chromatographically different from FAAH (Goparaju et al, 1999). Second, inhibition of FAAH activity in intact neurons and astrocytoma cells prevents the hydrolysis of anandamide, but has no effect on 2-AG degradation .…”

Section: -Ag Deactivation: Intracellular Hydrolysismentioning

confidence: 99%

The endogenous cannabinoid system and the treatment of marijuana dependence

Piomelli

2004

Neuropharmacology

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Enzymes of porcine brain hydrolyzing 2-arachidonoylglycerol, an endogenous ligand of cannabinoid receptors

Cited by 207 publications

References 41 publications

An optimized approach to study endocannabinoid signaling: evidence against constitutive activity of rat brain adenosine A₁ and cannabinoid CB₁ receptors

An optimized approach to study endocannabinoid signaling: evidence against constitutive activity of rat brain adenosine A₁ and cannabinoid CB₁ receptors

Chiral 3-(4,5-dihydrooxazol-2-yl)phenyl alkylcarbamates as novel FAAH inhibitors: Insight into FAAH enantioselectivity by molecular docking and interaction fields

The endogenous cannabinoid system and the treatment of marijuana dependence

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Enzymes of porcine brain hydrolyzing 2-arachidonoylglycerol, an endogenous ligand of cannabinoid receptors

Cited by 207 publications

References 41 publications

An optimized approach to study endocannabinoid signaling: evidence against constitutive activity of rat brain adenosine A1 and cannabinoid CB1 receptors

An optimized approach to study endocannabinoid signaling: evidence against constitutive activity of rat brain adenosine A1 and cannabinoid CB1 receptors

Chiral 3-(4,5-dihydrooxazol-2-yl)phenyl alkylcarbamates as novel FAAH inhibitors: Insight into FAAH enantioselectivity by molecular docking and interaction fields

The endogenous cannabinoid system and the treatment of marijuana dependence

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An optimized approach to study endocannabinoid signaling: evidence against constitutive activity of rat brain adenosine A₁ and cannabinoid CB₁ receptors

An optimized approach to study endocannabinoid signaling: evidence against constitutive activity of rat brain adenosine A₁ and cannabinoid CB₁ receptors