Biomedical subjects

Alexandros Makriyannis

Publications and source records attributed to Alexandros Makriyannis.

At least 19 recordsLinked to original sources

PubMed2007

The endogenous cannabinoid anandamide produces delta-9-tetrahydrocannabinol-like discriminative and neurochemical effects that are enhanced by inhibition of fatty acid amide hydrolase but not by inhibition of anandamide transport.

Anandamide is an endogenous ligand for brain cannabinoid CB(1) receptors, but its behavioral effects are difficult to measure due to rapid inactivation. Here we used a drug-discrimination procedure to test the hypothesis that anandamide, given i.v. or i.p., would produce discriminative effects like those of delta-9-tetrahydrocannabinol (THC) in rats when its metabolic inactivation was inhibited. We also used an in vivo microdialysis procedure to investigate the effects of anandamide, given i.v. or i.p., on dopamine levels in the nucleus accumbens shell in rats. When injected i.v., methanandamide (AM-356), a metabolically stable anandamide analog, produced clear dose-related THC-like discriminative effects, but anandamide produced THC-like discriminative effects only at a high 10-mg/kg dose that almost eliminated lever-press responding. Cyclohexyl carbamic acid 3'-carbamoyl-biphenyl-3-yl ester (URB-597), an inhibitor of fatty acid amide hydrolase (FAAH), the main enzyme responsible for metabolic inactivation of anandamide, produced no THC-like discriminative effects alone but dramatically potentiated discriminative effects of anandamide, with 3 mg/kg anandamide completely substituting for the THC training dose. URB-597 also potentiated the ability of anandamide to increase dopamine levels in the accumbens shell. The THC-like discriminative-stimulus effects of anandamide after URB-597 and methanandamide were blocked by the CB1 receptor antagonist rimonabant, but not the vanilloid VR1 receptor antagonist capsazepine. Surprisingly, the anandamide transport inhibitors N-(4-hydroxyphenyl)-eicosa-5,8,11,14-tetraenamide (AM-404) and N-(3-furylmethyl)eicosa-5,8,11,14-tetraenamide (UCM-707) did not potentiate THC-like discriminative effects of anandamide or its dopamine-elevating effects. Thus, anandamide has THC-like discriminative and neurochemical effects that are enhanced after treatment with a FAAH inhibitor but not after treatment with transport inhibitors, suggesting brain area specificity for FAAH versus transport/FAAH inactivation of anandamide.

Amidohydrolases↗

PubMed2006

Endocannabinoid metabolomics: a novel liquid chromatography-mass spectrometry reagent for fatty acid analysis.

We have synthesized 4,4-dimethoxyoxazoline derivatives of several fatty acids associated with the endocannabinoid metabolome using tris(hydroxymethyl)aminomethane in a 1-step reaction by microwave irradiation. The derivatization incorporates a nitrogen into the final product, which allows for improved detection by liquid chromatography-mass spectrometry in positive atmospheric pressure chemical ionization (APCI) mode. Palmitic and oleic acid derivatives show a 200-fold increase in sensitivity compared with the free acids when analyzed in negative-mode APCI. In addition to improving sensitivity, the oxazoline derivatization creates a similar ionization response for the fatty acids tested, which simplifies their quantitation. Fatty acid oxazoline derivatives can be detected using the same conditions optimized for the endocannabinoids, which allows for a simultaneous quantitation of the entire endocannabinoid metabolome.

Cannabinoid Receptor Modulators↗

PubMed2006

Discriminative stimulus functions of AM-1346, a CB1R selective anandamide analog in rats trained with Delta9-THC or (R)-methanandamide (AM-356).

OBJECTIVE: To characterize in vivo the high-affinity cannabinoid CB1 receptor (CB1R) selective anandamide analog AM-1346 [alkoxyacid amide of N-eicosa-tetraenylamine] using drug discrimination procedures. D-amphetamine and also morphine in the (R)-methanandamide-trained group (see below) were examined to assess pharmacological specificity. METHODS: Three groups of rats were trained to discriminate between vehicle and (1) 1.8 mg/kg Delta9-tetrahydrocannabinol (Delta9-THC); (2) 5.6 mg/kg Delta9-THC; and (3) 10 mg/kg (R)-methanandamide (AM-356; a metabolically stable analog of anandamide). Delta9-THC was given i.p. 30 min and (R)-methanandamide 15 min before training. RESULTS: AM-1346 generalized to all three training conditions, both at 15 and 30 min after administration. The rank order potency was: Delta9-THC > AM-1346 > (R)-methanandamide. AM-1346 appeared slightly more potent 30 min compared to 15 min postadministration. In the presence of 0.3 mg/kg of the CB1R antagonist/inverse agonist SR-141716A, the dose generalization curves of Delta9-THC and AM-1346 resulted in parallel shifts to the right in the 1.8 mg/kg Delta9-THC-trained group. A long duration of action for AM-1346 (relative to AM-356) was indicated in tests where AM-1346 was examined in the 5.6 mg/kg Delta9-THC-trained group. Neither D-amphetamine, nor morphine generalized in either of the groups, suggesting pharmacological specificity. CONCLUSION: Unlike (R)-methanandamide, the surmountable antagonism between SR-141716A and AM-1346 shows that the structural features of anandamide can be modified in ways that reduce the dissociation between the discriminative stimulus and rate decreasing effects of CB1R agonists derived from an anandamide template.

Analysis of Variance↗

PubMed2006

Synthesis and biological evaluation of benzopyran analogues bearing class III antiarrhythmic pharmacophores.

We have synthesized a series of compounds combining the hydroxy-benzopyran ring of vitamin E with the methylsulfonylaminophenyl group of class III antiarrhythmic drugs, connected through tertiary amine moieties. Evaluation of the antiarrhythmic and antioxidant activity of the new compounds was carried out on isolated rat heart preparations using the non-recirculating Langendorff mode. The new analogues were present, at 10 microM concentration, during ischemia and reperfusion. Selected compounds were further studied by a conventional microelectrode method in order to get insight into their cellular mode of action. The most active compound, N-[4-[2-[[2-(3,4-dihydro-6-hydroxy-2,2,7,8-tetramethyl-2H-1-benzopyran-5-yl)ethyl] methylamine]ethyl]phenyl]methanesulfonamide (19a), reduces premature beats, prolongs QT and QRS intervals during ischemia and reperfusion, and reduces MDA content, leading to a fast recovery of the heart. In addition, it exhibits moderate class III antiarrhythmic action.

Action Potentials↗

PubMed2006

AM1241, a cannabinoid CB2 receptor selective compound, delays disease progression in a mouse model of amyotrophic lateral sclerosis.

Effective treatment for amyotrophic lateral sclerosis (ALS) remains elusive. Motor neuron degeneration is the primary pathology in ALS; however non-neuronal cells contribute to the disease process. In particular, inflammatory processes have been shown to play an important role. AM1241 is a cannabinoid CB2 receptor selective agonist that has been shown to be effective in models of inflammation and hyperalgesia. Here we report that treatment with AM1241 was effective at slowing signs of disease progression when administered after onset of signs in an ALS mouse model (hSOD1(G93A) transgenic mice). Administration at the onset of tremors delayed motor impairment in treated mice when compared to vehicle controls. Three conditions of ALS, the loss of motor function, paralysis scoring and weight loss, were analyzed using a mathematical model. Loss of motor function (as assessed by performance on a rotarod) was delayed by 12.5 days in male mice by AM1241. In female mice, AM1241 extended rotarod performance by 3 days, although this was not statistically significant. In male mice, AM1241 also extended by 5 days the time to reach the 50% point on a visually-assessed performance scale. AM1241 did not affect weight loss or survival (129.8+/-1.7 days, vehicle; 129.1+/-7.0 days, AM1241, n=16). As AM1241 was well tolerated by the animals, cannabinoid CB2 receptor-selective compounds may be the basis for developing new drugs for the treatment of ALS and other chronic neurodegenerative diseases.

Amino Acid Substitution↗

PubMed2006

CB2 cannabinoid receptor mediation of antinociception.

Management of acute pain remains a significant clinical problem. In preclinical studies, CB2 cannabinoid receptor-selective agonists inhibit nociception without producing central nervous system side effects. The CB2 receptor-selective agonist AM1241 produces antinociceptive effects that are antagonized by CB2, but not CB1, receptor-selective antagonists, suggesting that activation of CB2 receptors results in antinociception. However, it has not been possible to definitively demonstrate that these effects are mediated by CB2 receptors, because we have lacked the pharmacological tools to confirm the in vivo receptor selectivity of the antagonists used. Further, recent evidence for cannabinoid-like receptors beyond CB1 and CB2 raises the possibility that AM1241 exerts its antinociceptive effects at uncharacterized CB2-like receptors that are also inhibited by AM630. The experiments reported here further test the hypothesis that CB2 receptor activation inhibits nociception. They evaluated the antinociceptive actions of AM1241 and the less-selective CB2 receptor agonist WIN55,212-2 in wild-type (CB2+/+) mice and in mice with genetic disruption of the CB2 receptor (CB2-/- mice). AM1241 inhibited thermal nociception in CB2+/+ mice, but had no effect in CB2-/- littermates. WIN55,212-2 produced equivalent antinociception in CB1+/+ and CB1-/- mice, while its antinociceptive effects were reduced in CB2-/- compared to CB2+/+ mice. The effects of morphine were not altered in CB2-/- compared to CB2+/+ mice. These data strongly suggest that AM1241 produces antinociception in vivo by activating CB2 cannabinoid receptors. Further, they confirm the potential therapeutic relevance of CB2 cannabinoid receptors for the treatment of acute pain.

Analgesics↗

PubMed2006

Suppression of food intake and food-reinforced behavior produced by the novel CB1 receptor antagonist/inverse agonist AM 1387.

Cannabinoid CB1 receptor antagonist/inverse agonists are becoming increasingly recognized for their potential therapeutic utility as appetite suppressants. In the current paper we characterize the biochemical and behavioral effects of AM 1387, which is a novel CB1 antagonist. AM 1387 exhibited binding affinity and selectivity for the CB1 over the CB2 receptor. Moreover, AM 1387 decreased GTPgammaS (EC50: 22.82 nM) and increased forskolin-stimulated cAMP (EC50: 274.6 nM), as did the CB1 inverse agonist AM 251 (GTPgammaS EC50: 25.82 nM; cAMP EC50: 363.8 nM), indicating that AM1387 also has inverse agonist properties in vitro. In the behavioral characterization in rats, AM 1387 suppressed lever pressing for food on two operant schedules (fixed-ratio 1 and 5). Timecourse of the effect on fixed-ratio 5 responding was then determined, and the half-life (t1/2=4.87 h) was found to be threefold shorter than what has been shown for SR 141716A, and fourfold shorter than AM 251. Finally, AM 1387 was found to suppress food intake using three diets of differing macronutrient composition and palatability. It was concluded that AM 1387 may be a useful tool for examining the effects of CB1 receptor antagonism or inverse agonism on food intake.

Animals↗

PubMed2006

3D-QSAR studies of arylpyrazole antagonists of cannabinoid receptor subtypes CB1 and CB2. A combined NMR and CoMFA approach.

The present work focuses on the study of the three-dimensional (3D) structural requirements for selective antagonist activity of arylpyrazole compounds at the cannabinoid CB1 and CB2 receptors. Initially, a combined high-resolution two-dimensional (2D) NMR and computer modeling approach was carried out to study the solution structure of the key pyrazole derivative N-(piperidin-1-yl)-5-phenyl-1-(n-pentyl)-4-methyl-1H-pyrazole-3-carboxamide (AM263). By using the NMR-determined molecular conformers as templates, the 3D quantitative structure-activity relationship (QSAR) studies were performed with the comparative molecular field analysis (CoMFA) approach on a set of arylpyrazole cannabinoid receptor antagonists. Molecular alignments suitable for deriving valuable pharmacophoric features for this series of compounds were determined. Such systematic 3D-QSAR/CoMFA analyses of 29 molecules and their receptor affinities gave guidance for understanding the binding affinities of arylpyrazoles at the CB1 and CB2 binding sites, respectively. Comparison of CoMFA steric and potential contour maps for affinity at the two cannabinoid receptor subtypes helps to differentiate structural requirements for each subtype and serves as a basis for the design of later-generation analogues.

Amino Acid Motifs↗

PubMed2006

Structural modifications of the cannabinoid side chain towards C3-aryl and 1',1'-cycloalkyl-1'-cyano cannabinoids.

The compounds reported in this study are Delta(8)-THC analogues in which the C3 five-carbon linear side chain of Delta(8)-THC was replaced with aryl and 1',1'-cycloalkyl substituents. Of the compounds described here analogues 2d (CB(1), K(i)=11.7 nM. CB(2), K(i)=9.39 nM) and 2f (CB(1), K(i)=8.26 nM. CB(2), K(i)=3.86 nM) exhibited enhanced binding affinities for CB(1) and CB(2), exceeding that of Delta(8)-THC. Efficient procedures for the synthesis of these novel cannabinoid analogues are described.

Animals↗

PubMed2006

Evaluation of the in vivo receptor occupancy for the behavioral effects of cannabinoids using a radiolabeled cannabinoid receptor agonist, R-[125/131I]AM2233.

G-protein coupled receptors exist in both high and low agonist affinity conformations, with tracer levels of agonist radioligands preferentially binding to the former. The goal of the present study was to characterize the in vivo binding of the aminoalkyindole-based, CB1 receptor agonist, R-[125/131I]AM2233 ((2-[125/131I]iodo-phenyl)-[1-(1-methyl-piperidin-2-yl-methyl)-1H-indol-3-yl]-methanone), and to use this radiotracer to selectively measure the receptor occupancy by the related CB1 receptor agonist, WIN55212-2, to the agonist-preferring affinity state of the receptor. In mouse locomotor assays, both WIN55212-2 and AM2233 (racemic) produced an approximately 60% reduction in activity at 1 mg/kg, (i.v.) and completely inhibited activity at 3 mg/kg, confirming their agonist nature. In ex vivo autoradiography, preferential uptake of R-[131I]AM2233 was apparent in CB1 receptor-rich areas, including globus pallidus, substantia nigra, striatum, cerebellum, and hippocampus. Overall brain uptake of R-[131I]AM2233 was 1.3% injected activity/g at 5 min in mice. Coinjection of 3 mg/kg (i.v.) SR141716A, a CB1 receptor antagonist, with R-[125I]AM2233 inhibited the radiotracer binding almost to nonspecific levels in the striatum, globus pallidus, and substantia nigra, although residual binding to a non-CB1 receptor remained in the hippocampus. In contrast to the effect of SR141716A, coinjection of 10 mg/kg (i.v.) WIN55212-2, a high dose that produced an immediate and profound immobility and catalepsy in the mice, reduced CB1 receptor-specific binding of R-[125I]AM2233 in CB1 receptor-rich areas by only 21-43%. These observations suggest that the behavioral effects of CB1 receptor agonists are manifested with a relatively small fraction of the agonist-preferring affinity state of the receptor occupied.

Animals↗

PubMed2006

Methods for the synthesis of cannabinergic ligands.

During the last decade, numerous cannabinergic ligands with high affinity and selectivity profiles for cannabinoid receptors (CB1 and CB2) emerged from rigorously pursued structure-activity relationship studies. This chapter focuses on the synthetic aspects of key cannabinoid receptor probes representing the different classes of cannabinergic ligands that encompasses classical cannabinoids (CCs) including some covalent binding derivatives, nonclassical cannabinoids (NCCs), hybrid cannabinoids, aminoalkylindoles (AAIs), diarylpyrazoles, and the endocannabinoids.

Cannabinoids↗

PubMed2006

Targeting the endocannabinoid system in treating brain disorders.

Recent cannabinoid research has a primary focus on developing therapeutics against human diseases. Many studies on cannabinoids indicate important progress for protection against several neurodegenerative disorders. Agonists of cannabinoid receptors activate signalling pathways in the brain that are linked to neuronal repair and cell maintenance, and endogenous ligands can also activate neuroprotective responses. These endocannabinoids are bioactive fatty acid amides and esters that are synthesised in the brain and include arachidonoyl ethanolamide (anandamide) and 2-arachidonoyl glycerol. Endocannabinoids are released in response to pathogenic events, thus representing a potential compensatory repair mechanism. Enhancing this on-demand action of endocannabinoids is a strategy with which to promote endogenous repair signalling. For such enhancement, considerable work has gone into modulating the availability of endocannabinoids by blocking the processes of their deactivation. The targets include the anandamide-hydrolysing enzyme fatty acid amide hydrolase, the carrier-mediated anandamide transport system and 2-arachidonoyl glycerol-deactivating enzyme monoacylglycerol lipase. The activity of endocannabinoids is terminated through transport and degradation and, accordingly, selective inhibitors of these processes effectively exploit the protective nature of cannabinergic responses. This review highlights recent studies implicating the endocannabinoid system in neuroprotection against different disorders of the CNS.

Animals↗

PubMed2006

Cannabinoid receptors as therapeutic targets.

The cannabinoid receptors CB1 and CB2 are family A, G-protein Coupled Receptors that mediate the effects of cannabinoids, a class of compounds that are so named because the first members were isolates of the cannabis plant. In recent history, there has been much anecdotal evidence that the potent and diverse physiological responses produced by these compounds can be turned to therapeutic benefit for a wide variety of maladies. The remarkable abundance of cannabinoid receptors and the discovery of several endogenous ligands along with enzyme and transporter proteins for which they are substrates, suggests that an endogenous cannabinoid neuromodulatory system is an important mediator of biological function. For these reasons CB1 and CB2 receptors are attractive targets for the design of therapeutic ligands. The action of these receptors, however, may also be modulated by manipulating the enzymes and membrane transporters that regulate the endogenous ligands. Despite the range of physiological processes and activities that are mediated by cannabinoid receptors, it is clear that it is possible to produce ligands that result in differential responses. In this paper, we review the pharmacophoric elements that lead to these differential responses and in order to discuss them in context we present an overview of structural aspects governing cannabinoid receptor function, the cannabinergic system and its physiological functions.

Animals↗

PubMed2006

The cannabinergic system as a target for anti-inflammatory therapies.

Habitual cannabis use has been shown to affect the human immune system, and recent advances in endocannabinoid research provide a basis for understanding these immunomodulatory effects. Cell-based experiments or in vivo animal testing suggest that regulation of the endocannabinoid circuitry can impact almost every major function associated with the immune system. These studies were assisted by the development of numerous novel molecules that exert their biological effects through the endocannabinoid system. Several of these compounds were tested for their effects on immune function, and the results suggest therapeutic opportunities for a variety of inflammatory diseases such as multiple sclerosis, rheumatoid arthritis, inflammatory bowel disease, atherosclerosis, allergic asthma, and autoimmune diabetes through modulation of the endocannabinoid system.

Animals↗

PubMed2005

Antagonism of discriminative stimulus effects of delta(9)-THC and (R)-methanandamide in rats.

RATIONALE: In previous drug discrimination studies we observed surmountable antagonism by Delta(9)-tetrahydrocannabinol (THC) in the presence of constant doses of SR-141716 [N-(piperidin-1-yl)-5-(4-chloro-phenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] (0.3 and 1 mg/kg), but there was only marginal evidence for surmountable antagonism with combinations of SR-141716 and (R)-methanandamide, a chiral analog of the endocannabioid anandamide. OBJECTIVE: Here we examine antagonism where the cannabinoid CB1 receptor agonist [Delta(9)-THC and (R)-methanandamide] dose is held constant (i.e., the training dose) and the antagonist {i.e., SR-141716 and AM-251 [N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide; 2 ml/kg]} dose varied. We also tested the cannabinoid CB2 receptor antagonist SR-144528 {N-[(1S)-endo-1,3,3-trimethylbicyclo(2.2.1)heptan-2-yl]5-(4-chloro-3-methyl-phenyl)-1-(4-methylbenzyl)pyrazole-3-carboxamide}. METHODS: Different groups of rats were trained to discriminate between vehicle and three different doses of Delta(9)-THC (1.8, 3, and 5.6 mg/kg, presumably reflecting different efficacy demands) as well as 10 mg/kg (R)-methanandamide. Dose-generalization tests involved different doses of the cannabinoid CB1 receptor agonists. Antagonist tests varied the dose of the antagonist (range: 0.1 and 3 mg/kg for SR-141716 and AM-251, and 1 to 10 mg/kg for SR-144528). RESULTS: SR-141716 and AM-251 doses dependently blocked the agonist-induced discriminative stimulus effects. SR-141716 tended to be slightly more potent than AM-251. The effective dose 50 (ED(50)) of SR-141716 was higher in the 5.6 mg/kg Delta(9)-THC-trained group relative to the two other Delta(9)-THC-trained groups. The cannabinoid CB2 receptor antagonist SR-144528 combined with the training dose of 1.8 mg/kg Delta(9)-THC, as well as when combined with the training dose of 10 mg/kg (R)-methanandamide, did not markedly change drug-appropriate (agonist) responses. CONCLUSION: Data support that the discriminative stimulus effects of (R)-methanandamide and its overlap with the Delta(9)-THC cue are, indeed, CB1 receptor mediated events as revealed in antagonism tests with the selective central CB1 receptor antagonists SR-141716 and AM-251. The activation of cannabinoid CB2 receptors appears to be insignificant for these discriminations.

Animals↗

PubMed2005

Natural cannabinoids: templates for drug discovery.

Recent studies have elucidated the biosynthetic pathway of cannabinoids and have highlighted the preference for a C-3 n-pentyl side chain in the most prominently represented cannabinoids from Cannabis sativa and their medicinally important decarboxylation products. The corresponding C-3 n-propyl side chain containing cannabinoids are also found, although in lesser quantities. Structure-activity relationship (SAR) studies performed on Delta(9)-tetrahydrocannabinol (Delta(9)-THC), the key psychoactive ingredient of Cannabis, and its synthetic analogues have identified the C-3 side chain as the key pharmacophore for ligand affinity and selectivity for the known cannabinoid receptors and for pharmacological potency. Interestingly, the terminal n-pentyl saturated hydrocarbon side chain of endocannabinoids also plays a corresponding crucial role in conferring similar properties. This review briefly summarizes the biosynthesis of cannabinoids and endocannabinoids and focuses on their side chain SAR.

Animals↗

PubMed2005

Identification and functional characterization of brainstem cannabinoid CB2 receptors.

The presence and function of CB2 receptors in central nervous system (CNS) neurons are controversial. We report the expression of CB2 receptor messenger RNA and protein localization on brainstem neurons. These functional CB2 receptors in the brainstem were activated by a CB2 receptor agonist, 2-arachidonoylglycerol, and by elevated endogenous levels of endocannabinoids, which also act at CB1 receptors. CB2 receptors represent an alternative site of action of endocannabinoids that opens the possibility of nonpsychotropic therapeutic interventions using enhanced endocannabinoid levels in localized brain areas.

Animals↗

PubMed2005

Potent cannabinergic indole analogues as radioiodinatable brain imaging agents for the CB1 cannabinoid receptor.

A series of novel aminoalkylindoles was synthesized in an effort to develop compounds that are potent agonists at the CB1 cannabinoid receptor and that are also easily labeled with radioisotopes of iodine for biochemical and imaging studies. 2-Iodophenyl-[1-(1-methylpiperidin-2-ylmethyl)-1H-indol-3-yl]methanone (8, AM2233) had a very high affinity for the rat CB1 receptor, with most of the affinity residing with the (R)-enantiomer. Radioiodinated 8, (R)-8, and (S)-8 were prepared by radioiododestannylation of the tributyltin analogues in high yields, radiochemical purities, and specific radioactivities. In a mouse hippocampal membrane preparation with [131I](R)-8 as radioligand, racemic 8 exhibited a K(i) value of 0.2 nM compared with 1.6 nM for WIN55212-2. In autoradiographic experiments with mouse brain sections, the distribution of radioiodinated 8 was consistent with that of brain CB1 receptors. Again, very little specific binding was seen with the (S)-enantiomer [131I](S)-8 and none occurred with the (R)-enantiomer [131I](R)-8 in sections from CB1 receptor knockout mice. Radioiodinated 8 thus appears to be a suitable radioligand for studies of CB1 cannabinoid receptors.

Animals↗