Design, synthesis and evaluation of resveratrol-indazole hybrids as novel monoamine oxidases inhibitors with amyloid-ß aggregation inhibition.

Novel hybrids with MAO and Aß (1-42) self-aggregation inhibitory activities were designed and synthesized with the employment of indazole moiety and resveratrol. The biological screening results indicated that most compounds displayed potent inhibitory activity for Aß (1-42) self-aggregation, and obvious selective inhibition to MAO-B. Among these compounds, compound 6e was the most potent inhibitor not only for hMAO-B (IC50 = 1.14 µM) but also for Aß (1-42) self-aggregation (58.9% at 20 µM). Molecular modeling and kinetic studies revealed that compound 6e was a competitive MAO-B inhibitor, which can occupy the active site of MAO-B, and interact with Aß (1-42) via π-π and cation-π stacking interactions. In addition, compound 6e had no toxicity on PC12 cells and could cross the BBB. Collectively, all these results suggested that compound 6e might be a promising multi-target lead compound worthy of further investigation.

Characterization of 3-iodothyronamine in vitro dynamics by mathematical modeling.

3-Iodothyronamine (T1AM) is regarded as a hormone-like substance thanks to its endogenous nature, its interaction with specific receptors trace amine-associated receptor 1 and its biological effects. We characterized T1AM transport and conversion in an in vitro culture of H9c2 murine cells, after a T1AM bolus injection. Samples of cell medium culture and cell lysate were assayed by high-performance liquid chromatography coupled to tandem mass spectrometry. We performed comparative experiments by adding to T1AM bolus amino oxidase inhibitors as iproniazid, pargyline (monoamine oxidase, MAO inhibitors), aminoguanidine, and semicarbazide (semicarbazide-sensitive amino oxidase, SSAO inhibitors). A mathematical model was developed, based on the assumption that T1AM is transported with a mechanism that is typical of hormone transport (i.e., EGF or insulin). We noticed that surface receptors downregulation could play a major role in T1AM dynamics. We also estimated that T1AM catabolism is mainly affected by MAO inhibitors, which produce a dramatic decrease in the kinetic constants related to T1AM degradation, while no significant changes were observed in experiments with SSAO inhibitors.

MAO inhibitory activity modulation: 3-Phenylcoumarins versus 3-benzoylcoumarins.

With the aim of finding the structural features for the human MAO inhibitory activity and selectivity, in the present communication we report the synthesis, pharmacological evaluation and a comparative study of a new series of 3-phenylcoumarins (compounds 1-4) and 3-benzoylcoumarins (compounds 5-8). A bromo atom and a methoxy/hydroxy substituent were introduced in these scaffolds, at six and eight positions of the coumarin moiety, respectively. The synthesized compounds 1-8 were evaluated as MAO-A and B inhibitors using R-(-)-deprenyl and iproniazide as reference compounds. The presence or absence of a carbonyl group between the coumarin and the phenyl substituent in 3 position remarks, respectively, the MAO-A or MAO-B inhibitory activity. Some of the new compounds showed MAO-B inhibitory activities in the low nanomolar range. Compound 2 (IC(50)=1.35nM) showed higher inhibitory activity than the R-(-)-deprenyl (IC(50)=19.60nM) and higher MAO-B selectivity, with more than 74,074-fold inhibition level, respecting to the MAO-A isoform.

Inhibition of monoamine oxidase activity by cannabinoids.

Brain monoamines are involved in many of the same processes affected by neuropsychiatric disorders and psychotropic drugs, including cannabinoids. This study investigated in vitro effects of cannabinoids on the activity of monoamine oxidase (MAO), the enzyme responsible for metabolism of monoamine neurotransmitters and affecting brain development and function. The effects of the phytocannabinoid Delta(9)-tetrahydrocannabinol (THC), the endocannabinoid anandamide (N-arachidonoylethanolamide [AEA]), and the synthetic cannabinoid receptor agonist WIN 55,212-2 (WIN) on the activity of MAO were measured in a crude mitochondrial fraction isolated from pig brain cortex. Monoamine oxidase activity was inhibited by the cannabinoids; however, higher half maximal inhibitory concentrations (IC(50)) of cannabinoids were required compared to the known MAO inhibitor iproniazid. The IC(50) was 24.7 micromol/l for THC, 751 micromol/l for AEA, and 17.9 micromol/l for WIN when serotonin was used as substrate (MAO-A), and 22.6 micromol/l for THC, 1,668 micromol/l for AEA, and 21.2 micromol/l for WIN when phenylethylamine was used as substrate (MAO-B). The inhibition of MAOs by THC was noncompetitive. N-Arachidonoylethanolamide was a competitive inhibitor of MAO-A and a noncompetitive inhibitor of MAO-B. WIN was a noncompetitive inhibitor of MAO-A and an uncompetitive inhibitor of MAO-B. Monoamine oxidase activity is affected by cannabinoids at relatively high drug concentrations, and this effect is inhibitory. Decrease of MAO activity may play a role in some effects of cannabinoids on serotonergic, noradrenergic, and dopaminergic neurotransmission.

Inhibition of monoamine oxidase activity by antidepressants and mood stabilizers.

OBJECTIVE: Monoamine oxidase (MAO), the enzyme responsible for metabolism of monoamine neurotransmitters, has an important role in the brain development and function, and MAO inhibitors have a range of potential therapeutic uses. We investigated systematically in vitro effects of pharmacologically different antidepressants and mood stabilizers on MAO activity. METHODS: Effects of drugs on the activity of MAO were measured in crude mitochondrial fraction isolated from cortex of pig brain, when radiolabeled serotonin (for MAO-A) or phenylethylamine (for MAO-B) was used as substrate. The several antidepressants and mood stabilizers were compared with effects of well known MAO inhibitors such as moclobemide, iproniazid, pargyline, and clorgyline. RESULTS: In general, the effect of tested drugs was found to be inhibitory. The half maximal inhibitory concentration, parameters of enzyme kinetic, and mechanism of inhibition were determined. MAO-A was inhibited by the following drugs: pargyline > clorgyline > iproniazid > fluoxetine > desipramine > amitriptyline > imipramine > citalopram > venlafaxine > reboxetine > olanzapine > mirtazapine > tianeptine > moclobemide, cocaine >> lithium, valproate. MAO-B was inhibited by the following drugs: pargyline > clorgyline > iproniazid > fluoxetine > venlafaxine > amitriptyline > olanzapine > citalopram > desipramine > reboxetine > imipramine > tianeptine > mirtazapine, cocaine >> moclobemide, lithium, valproate. The mechanism of inhibition of MAOs by several antidepressants was found various. CONCLUSIONS: It was concluded that MAO activity is acutely affected by pharmacologically different antidepressants at relatively high drug concentrations; this effect is inhibitory. There are differences both in inhibitory potency and in mechanism of inhibition between both several drugs and the two MAO isoforms. While MAO inhibition is not primary biochemical effect related to their therapeutic action, it can be supposed that decrease of MAO activity may be concerned in some effects of these drugs on serotonergic, noradrenergic, and dopaminergic neurotransmission.

Metabolism of dopamine by the nasal mucosa.

The nasal route of administration offers several advantages over oral and intravenous administration, including the ability to avoid hepatic first pass metabolism. Dopamine deficiency has been associated with several neurological disorders; it has been shown to have good systemic bioavailability and significant uptake into the CNS following intranasal administration. The purpose of these studies was to investigate the limiting role of mucosal metabolism of dopamine during nasal absorption. In vitro transport and initial rate studies were carried out using nasal mucosal explants to study dopamine permeability and metabolism. Dihydroxyphenylacetic acid (DOPAC) was the only metabolite detected. Monoamine oxidase (MAO), the enzyme responsible for DOPAC formation, was localized to the submucosal region of the nasal explants. The amount of DOPAC formed during the transport studies was less than 0.5% of the initial amount of dopamine placed into the system. Iproniazid, an MAO inhibitor, blocked DOPAC formation but had no effect on dopamine transport. The limited extent of dopamine metabolism compared to its mucosal transport demonstrates that nasal dopamine transport is not significantly reduced by mucosal metabolism and suggests that the nasal route may be promising for the efficient delivery of dopamine to the CNS.

(-)-Trans-epsilon-viniferin, a polyphenol present in wines, is an inhibitor of noradrenaline and 5-hydroxytryptamine uptake and of monoamine oxidase activity.

(-)-Trans-epsilon-viniferin (epsilon-viniferin, 5-200 microM), a dimer of resveratrol, concentration-dependently inhibited the uptake of [3H]noradrenaline and [3H]5-HT by synaptosomes from rat brain (being slightly but significantly more selective against [3H]noradrenaline) and the uptake of [3H]5-HT by human platelets. On the other hand, epsilon-viniferin (5-200 microM) concentration-dependently inhibited the enzymatic activity of commercial (human recombinant) monoamine oxidase (MAO) isoform (MAO-A and MAO-B) activity, being slightly but significantly more selective against MAO-B than against MAO-A. Taking into account that the principal groups of drugs used to treat major depression are noradrenaline/5-HT uptake or MAO inhibitors, under the assumption that epsilon-viniferin exhibits a similar behaviour in humans in vivo, our results suggest that this natural polyphenol may be of value as a structural template for the design and development of new antidepressant drugs with two important biochemical activities combined in the same chemical structure: noradrenaline/5-HT uptake and MAO inhibitory activity.

A possible indirect sympathomimetic action of metformin in the arterial vessel wall of spontanously hypertensive rats.

The antidiabetic drug metformin (MF) typically achieves only micromolar levels in plasma with normal therapeutic use. However, it is also known to accumulate in various tissues up to several times higher after standard oral dosing and we now have evidence from both in vivo and in vitro experiments with spontaneously hypertensive rats (SHR) that millimolar levels stimulate release of norepinephrine (NE) from vascular sympathetic nerve endings (SNEs). As shown in the present work with SHR tail arterial tissue (rich in SNEs), the known vasodilator effect of millimolar levels of MF on the smooth muscle (even if contracted with a nonadrenergic agonist), is attenuated by the presence of the SNEs unless phentolamine (an alpha receptor blocker) is present. We reasoned that the mechanism for this apparent NE-releasing action of MF is not exocytotic release as that would require depolarization of the neuronal cell membranes in SNEs, and MF at millimolar levels is known to repolarize (not depolarize) membranes of other cells. Thus, we tested the possibility that MF releases NE by an indirect sympathomimetic-like action. Such an action should be amplified by monoamine oxidase inhibitors (e.g. iproniazid) and blocked by NE-carrier inhibitors (e.g. desipramine). Accordingly, we found that the abovementioned attenuating effect of intact SNEs on MF's relaxation of SHR tail arterial tissue (compared to tissues in which SNEs were experimentally removed with 6-hydroxydopamine) was amplified nearly 3-fold by iproniazid (p<0.05) and blocked by desipramine (p<0.05). These results support an indirect sympathomimetic action of MF and raise the question whether commonly used antidepressants with properties similar to iproniazid and desipramine might alter MF's beneficial vasodilatory (and thus antihypertensive) effectiveness in diabetic patients with hypertension.

Inhibition of type A monoamine oxidase by coptisine in mouse brain.

The inhibitory effects of coptisine, a protoberberine isoquinoline alkaloid, on type A and type B monoamine oxidase (MAO-A and MAO-B) activities in mouse brain were investigated. Coptisine showed an inhibitory effect on MAO-A activity in a concentration-dependent manner using a substrate kynuramine, but coptisine did not inhibit MAO-B activity. Coptisine exhibited 54.3% inhibition of MAO-A activity at 2 microM. The values of Km and Vmax of MAO-A were 151.9 +/- 0.6 microM and 0.40 +/- 0.03 nmol/min/mg protein, respectively (n=5). Coptisine competitively inhibited MAO-A activity with kynuramine. The Ki value of coptisine was 3.3 microM. The inhibition of MAO-A by coptisine was found to be reversible by dialysis of the incubation mixture. These results suggest that coptisine is a potent reversible inhibitor of MAO-A, and that coptisine functions to regulate the catecholamine content.

Characterization of phentermine and related compounds as monoamine oxidase (MAO) inhibitors.

Phentermine was shown in the 1970s to inhibit the metabolism of serotonin by monoamine oxidase (MAO), but never was labeled as an MAO inhibitor; hence, it was widely used in combination with fenfluramine, and continues to be used, in violation of their labels, with other serotonin uptake blockers. We examined the effects of phentermine and several other unlabeled MAO inhibitors on MAO activities in rat lung, brain, and liver, and also the interactions of such drugs when administered together. Rat tissues were assayed for MAO-A and -B, using serotonin and beta-phenylethylamine as substrates. Phentermine inhibited serotonin-metabolizing (MAO-A) activity in all three tissues with K(i) values of 85-88 microM. These potencies were similar to those of the antidepressant MAO inhibitors iproniazid and moclobemide. When phentermine was mixed with other unlabeled reversible MAO inhibitors (e.g. pseudoephedrine, ephedrine, norephedrine; estradiol benzoate), the degree of MAO inhibition was additive. The cardiac valvular lesions and primary pulmonary hypertension that have been reported to be associated with fenfluramine-phentermine use may have resulted from the intermittent concurrent blockage of both serotonin uptake and metabolism.

Evidence that histamine is involved as a mediator of endothelium-dependent contraction induced by A23187 in bovine intrapulmonary vein.

This study was initiated to test the hypothesis that histamine can act as an endothelium-derived contracting factor in bovine isolated intrapulmonary vein. The effects of calcium ionophore, calcimycin (A23187), on isometric tension were compared in unstimulated rings of intrapulmonary vein with and without endothelium. A23187 (0.1-10 microM) induced concentration-related contraction when endothelium was present. Destruction of endothelium markedly inhibited A23187-induced contraction. Methylene blue, hemoglobin or NG-methyl-L-arginine significantly enhanced A23187-induced contraction only in venous rings with endothelium consistent with attenuation of the contraction by the concomitant release of endothelium-derived relaxing factor (nitric oxide) [EDRF(NO)]. Histamine H1 receptor antagonists inhibited, and iproniazid enhanced, contraction elicited by A23187. A23187 induced release of greater amounts of histamine from venous rings with than without endothelium. A23187-induced contraction was not mimicked by the mast cell activator, compound 48/80, and was not inhibited by preexposure to compound 48/80 or in the presence of cromolyn or doxantrazole. A23187-induced contraction was not inhibited by pretreatment with indomethacin, phentolamine, lipoxygenase inhibitors or superoxide dismutase. The results indicate that A23187 induces endothelium-dependent contraction in bovine intrapulmonary vein and support histamine as one major mediator involved. The association of destruction of endothelium with an inhibition of both A23187-induced contraction and histamine release is consistent with the endothelium as a source for histamine which can exert a local vasoconstrictor effect in bovine intrapulmonary vein.

Acrolein genotoxicity in Drosophila melanogaster. III. Effects of metabolism modification.

In order to investigate the role of metabolism in acrolein genotoxicity in D. melanogaster, the action of several metabolism modifiers, namely phenobarbital, an inducer of xenobiotic metabolism, phenylimidazole and iproniazid, inhibitors of oxidative activities of cytochrome P450, and diethyl maleate, a glutathione-depleting agent, have been assayed using the sex-linked recessive lethal (SLRL) test, with two different administration routes (feeding and injection). The results support the hypothesis that acrolein is not only a direct mutagen but is also transformed, by oxidative activities of cytochrome P450 after glutathione conjugation, into an active metabolite, possibly glycidaldehyde. Moreover, acrolein is deactivated by an enzymatic activity induced by phenobarbital.

Catabolism of 5-aminolevulinic acid to CO2 by rat liver mitochondria.

5-Aminolevulinic acid (ALA), the heme precursor accumulated in plasma and several organs of carriers of acute intermittent porphyria, hereditary tyrosinemia, and saturnism, was previously shown to yield reactive oxygen species upon metal-catalyzed aerobic oxidation and to cause the in vivo and in vitro impairment of rat liver mitochondrial functions. We have studied the uptake and catabolism of [5-14C]ALA to CO2 by isolated rat liver mitochondria (RLM) with the aim of determining whether possible ALA-driven oxidative injury to mitochondria can also occur into the matrix. Using silicone oil centrifugation of [5-14C]ALA-treated RLM, ALA was found to partition evenly into the intra- and extramatrix space of the mitochondrial preparations. The yield of evolved 14CO2 is very low (0.2%), responds to the concentration of added ADP, and is inhibited by malonate (75% at 2 mM), iproniazid (45% at 2 mM), beta-chloroalanine (36% at 1 mM), and aminooxyacetate (55% at 0.1 mM). With both iproniazid and aminooxyacetate, the percentage of inhibition is the same as that observed with the latter inhibitor alone. These data indicate that ALA decarboxylation by the Krebs cycle is a minor process and that it is initiated enzymically (transaminase) and not by metal-catalyzed ALA autoxidation.

Effects of two substituted hydrazine monoamine oxidase (MAO) inhibitors on neurotransmitter amines, gamma-aminobutyric acid, and alanine in rat brain.

Time- and dose-response analyses were undertaken to investigate the effects of the substituted hydrazine monoamine oxidase (MAO) inhibitors iproniazid and nialamide on the following: MAO-A and -B activity; levels of gamma-aminobutyric acid (GABA), alanine (ALA), and the neurotransmitter amines dopamine, noradrenaline, and 5-hydroxytryptamine (serotonin) and their acid metabolites; and the activity of GABA-transaminase and ALA-transaminase. The results showed that these drugs are relatively potent MAO inhibitors but, unlike the unsubstituted hydrazine MAO inhibitor phenelzine, they do not produce increased GABA and ALA levels in brain. These experiments suggest that a free hydrazine group is necessary for MAO inhibitors to also have marked effects on GABA and ALA.

Evidence against serotonin-induced endothelium-dependent relaxation in bovine coronary artery.

Bovine coronary artery rings were mounted in tissue baths for the measurement of isometric contraction in order to test for serotonin-induced endothelium-dependent relaxation. A23187 caused a concentration-dependent relaxation in precontracted artery rings when the endothelium was intact, but not when it was removed. Endothelium removal had no effect on serotonin concentration contractile response curves (CRC) in normal Krebs' solution, but enhanced the response to serotonin in artery rings precontracted with 25 mM K+ Krebs' solution. In endothelium-intact, precontracted artery rings, ketanserin shifted the serotonin CRC to the right, but did not convert the contraction to relaxation. 5-Carboxamidotryptamine caused a weak contraction that was blocked by ketanserin, but not converted to relaxation. Inactivation of monoamine oxidase enhanced the contractile response to serotonin in precontracted artery rings in a manner identical to that caused by endothelium removal. Inactivation of monoamine oxidase had no effect in endothelium-denuded artery rings. De-endothelialized rabbit aorta strips pretreated with phenoxybenzamine were placed in close apposition to endothelium-intact coronary artery strips. Superoxide dismutase, acetylcholine and A23187 caused the precontracted rabbit aorta strip to relax, whereas serotonin had no effect. These results support the conclusion that the endothelium of the bovine coronary artery releases a relaxing factor both spontaneously and in response to acetylcholine and A23187. Serotonin appears to be incapable of releasing endothelium-derived relaxing factor or causing an endothelium-dependent relaxation. However, bovine coronary artery endothelium appears to be able to take up and metabolize serotonin.

Dual effect of serotonin on growth of bovine pulmonary artery smooth muscle cells in culture.

We have previously reported that serotonin (5-hydroxytryptamine [5HT]) alters cultured bovine pulmonary artery smooth muscle cell (SMC) configuration through two different regulatory mechanisms. We now report that 5HT also regulates SMC growth through these same two mechanisms--a stimulatory event initiated intracellularly and inhibition of growth resulting from a cell surface action. 5HT (1 microM) plus 0.1 mM iproniazid (a 5HT metabolic inhibitor) produced a severalfold stimulation of DNA synthesis (as measured by [3H]thymidine incorporation) of SMCs after a 17-24-hour incubation with only a slight elevation of cellular cAMP. This stimulatory effect responded synergistically with other growth factors including platelet-derived growth factor, fibroblast growth factor, and epidermal growth factor and was effectively reversed by 5HT uptake inhibition. It was not produced by 5-hydroxyindoleacetic acid, a metabolite of 5HT. In the presence of 1 microM 5HT plus 0.1 mM isobutylmethylxanthine (IBMX), cAMP was elevated eightfold, dendritic formation occurred, and [3H]thymidine labeling of SMCs was inhibited. Inhibition of labeling by [3H]thymidine was mimicked by other agents that elevated cellular cAMP (10 microM histamine, 1 microM isoproterenol plus 0.1 mM IBMX, and 10 microM forskolin) and by 1 mM dibutyryl cAMP. This inhibitory effect was not blocked by either inhibition of 5HT uptake or 5HT-receptor antagonists ketanserin (5HT2); methiothepin, spiperone, and mianserin (5HT1/5HT2); and 3-tropanyl-indole-3-carboxylate and 3-tropanyl-3,5-dichlorobenzoate (5HT3). However, similar to 5HT, the 5HT1A agonist, (+/-)-8-hydroxy-(+/-)-2-dipropylamino-8-hydroxy-1,2,3, 4-tetrahydronaphthalenehydrobromide, in association with IBMX, produced an elevation in cAMP and inhibition of labeling by [3H]thymidine. 5HT, in the presence of either iproniazid or IBMX, did not alter [Ca2+]i, indicating that [Ca2+]i was not a signal for either of these actions.(ABSTRACT TRUNCATED AT 250 WORDS)

Use of pig hepatocytes to study the inhibition of monoamine oxidase by furazolidone.

Primary cultures of pig hepatocytes were used to examine the irreversible inhibition of monoamine oxidase (MAO), which has been observed in tissues of a number of different animal species after oral treatment with furazolidone. The rapid biotransformation of the MAO substrate p-tyramine by intact cells could effectively and irreversibly be inhibited with the known MAO inhibitors iproniazid and clorgyline. Incubation of cells with beta-hydroxyethylhydrazine and also 3-amino-2-oxazolidinone, which were previously proposed as the metabolites of furazolidone responsible for the in vivo effect, resulted in an irreversible inhibition of the MAO activity. Incubation of cells with furazolidone also resulted in a dose-related inhibition, but this effect was completely reversible on withdrawal of the drug. A similar MAO inhibition was observed after treatment of cells with nitrofurazone and furaltadone but not with nitrofurantoin. The results obtained with intact cells were confirmed by studies with 13,000 g pellets of homogenates made from cells preincubated for 24 hr with the compounds, which showed an irreversible inhibition in the case of iproniazid and 3-amino-2-oxazolidinone, but not in the case of furazolidone. The present study shows that hepatocytes are capable of transforming 3-amino-2-oxazolidinone, but not furazolidone itself, into a potent irreversible type of MAO inhibitor.

Effect of postnatal anoxia on seizure susceptibility in rats: bicuculline-induced seizures and pretreatment with iproniazid.

The effects of anoxia at 10 days of age on seizures induced by bicuculline (BIC), a specific antagonist of gamma-aminobutyric acid (GABA), and BIC-induced seizures after pretreatment with iproniazid (IPR), a monoamine oxidase inhibitor, were investigated in male adult rats. The rats with postnatal anoxia (Group A) were significantly more susceptible to BIC than the control rats (Group C). On the other hand, BIC-induced seizures in both groups were suppressed by IPR pretreatment. Especially, the suppression of BIC-induced seizures which was made by pretreatment with IPR at small doses was more apparent in Group A as compared with Group C. The present study suggests that the enhanced seizure susceptibility of adult rats with postnatal anoxia involves brain monoaminergic system.

Metabolism of diltiazem. III. Oxidative deamination of diltiazem in rat liver microsomes.

The main metabolites of diltiazem in rats are acidic metabolites having a carboxyl group which may be formed by oxidative deamination of the dimethylaminoethyl group of diltiazem. In order to identify the enzymes responsible for the deamination and formation of acidic and neutral metabolites [14C]diltiazem was incubated with microsomal and mitochondrial preparations from the liver of SD male rats. Both acidic and neutral metabolites were formed only in the presence of an nicotinamide adenine dinucleotide phosphate generating system. Their formation was remarkable, especially in the microsomes, and inhibited by SKF 252-A, but not by pargyline and iproniazid. The production of neutral metabolites surpassed that of acidic ones. Structural analysis by gas chromatography-mass spectrometry showed that the neutral metabolites are aldehydes which have not been detected in vivo. The results suggest that the dimethylaminoethyl group of diltiazem is oxidized to an aldehyde group by microsomal cytochrome P-450 in the liver. Subsequently, the aldehyde group would be dehydrogenated to the carboxyl group.