Selective inhibition of monoamine oxidase in monoaminergic neurons in the rat brain

The prevention by six reversible and selective monoamine oxidase-A (MAO-A) inhibitors (alpha-ethyl-tryptamine, harmaline, 4-methoxyamphetamine, amiflamine [FLA 336(+)], N-desmethylamiflamine [FLA 788(+)] and N,N-desmethylamiflamine [FLA 668(+)] of the phenelzine-induced irreversible MAO inhibition in the rat brain was examined. By using crude synaptosome preparations of hypothalamus and striatum incubated with low substrate concentrations of 14C-serotonin (1 X 10(-7) M), 14C-noradrenaline (2.5 X 10(-7) M) and 14C-dopamine (2.5 X 10(-7) M) in the absence and presence of selective amine uptake inhibitors (alaproclate, maprotiline and amfonelic acid, respectively), it was possible to determine the deaminating activities inside and outside the specific aminergic synaptosomes. Thus, with this technique the protection of MAO by the reversible inhibitors administered orally 1 h prior to the subcutaneous injection of phenelzine against the phenelzine effect could be determined inside and outside the specific aminergic neurons. It was found that alpha-ethyltryptamine, 4-methoxyamphetamine and particularly amiflamine and FLA 788(+) were more potent inside than outside the serotonergic neurons. FLA 668(+) was a selective inhibitor of noradrenergic MAO, to which also 4-methoxyamphetamine, amiflamine and FLA 788(+), but not alpha-ethyltryptamine had some preference. Harmaline had no certain preference for MAO in any of the aminergic neurons. At high doses of FLA 668(+) a preference for dopaminergic MAO was observed. Since pretreatment of the rats with norzimeldine or desipramine antagonized the preferences for serotonergic or noradrenergic MAO, it is plausible to conclude that the compounds showing these preferences are accumulated in the neurons by the membranal uptake systems.     

Inhibition of monoamine oxidase A-form and semicarbazide-sensitive amine oxidase by selective and reversible monoamine oxidase-A inhibitors, amiflamine and FLA 788(+)

In vitro studies demonstrated that two selective monoamine oxidase (MAO)-A inhibitors, amiflamine and FLA 788(+), have been shown to inhibit semicarbazide-sensitive amine oxidase (SSAO) in rat testis and lung homogenates in a concentration-dependent way. The inhibition was not greatly influenced by pretreatment of the preparations with either clorgyline (10(-3) mol/l), l-deprenyl (10(-3) mol/l) or SKF 525A (10(-4) mol/l). The two compounds showed a time-dependent inhibition of SSAO, and for the initial phase of the inhibition, amiflamine is a competitive inhibitor with a Kislope of 135 mumol/l, but FLA 788(+) is a noncompetitive inhibitor with a Ki value of 180 mumol/l. After preincubation for 60 min at 37 degrees C, however, inhibition by amiflamine was found to be essentially irreversible whereas that produced by FLA 788(+) was still noncompetitive and reversible. These two compounds also reversibly and competitively inhibited rat testis MAO-A with FLA 788(+) being much more selective towards this MAO (Kislope = 0.26 mumol/l for FLA 788(+) and 7 mumol/l for amiflamine, respectively). The present results indicate that both MAO-A-selective inhibitors also inhibit SSAO in vitro, but their properties as SSAO inhibitors differ from those as MAO-A inhibitors.     

Modification of blood pressure and nictitating membrane response to sympathetic amines by selective monoamine oxidase inhibitors, types A and B, in the cat.

The selective monoamine oxidase (MAO) inhibitors clorgyline, selegiline and AGN 1135 did not cause a change in responses of the cat nictitating membrane to preganglionic sympathetic nerve stimulation at 5 Hz. Both selective MAO-A and MAO-B inhibitors markedly potentiated nictitating membrane contractions in response to beta-phenylethylamine (PEA). However, the responses to tyramine were unchanged. The pressor responses to tyramine were potentiated by the selective MAO-A inhibitor clorgyline (2 mg kg-1) but not by selegiline (1.0 mg kg-1) and AGN 1135 (1.5 mg kg-1), selective MAO-B inhibitors. At the doses used selegiline and AGN 1135 caused a near total selective inhibition of liver and brain MAO-B, while clorgyline inhibited MAO-A only in the brain. AGN 1135, like selegiline, could be a useful drug in potentiating the action of L-DOPA in Parkinson's disease.     

Rasagiline [N-propargyl-1R(+)-aminoindan], a selective and potent inhibitor of mitochondrial monoamine oxidase B

1. Rasagiline [N-propargyl-1R(+)-aminoindan], was examined for its monoamine oxidase (MAO) A and B inhibitor activities in rats together with its S(-)-enantiomer (TVP 1022) and the racemic compound (AGN-1135) and compared to selegiline (1-deprenyl). The tissues that were studied for MAO inhibition were the brain, liver and small intestine. 2. While rasagiline and AGN1135 are highly potent selective irreversible inhibitors of MAO in vitro and in vivo, the S(-) enantiomer is relatively inactive in the tissues examined. 3. The in vitro IC(50) values for inhibition of rat brain MAO activity by rasagiline are 4.43+/-0.92 nM (type B), and 412+/-123 nM (type A). The ED(50) values for ex vivo inhibition of MAO in the brain and liver by a single dose of rasagiline are 0.1+/-0.01, 0.042+/-0.0045 mg kg(-1) respectively for MAO-B, and 6.48+/-0.81, 2.38+/-0.35 mg kg(-1) respectively for MAO-A. 4. Selective MAO-B inhibition in the liver and brain was maintained on chronic (21 days) oral dosage with ED(50) values of 0.014+/-0.002 and 0.013+/-0.001 mg kg(-1) respectively. 5. The degree of selectivity of rasagiline for inhibition of MAO-B as opposed to MAO-A was similar to that of selegiline. Rasagiline was three to 15 times more potent than selegiline for inhibition of MAO-B in rat brain and liver in vivo on acute and chronic administration, but had similar potency in vitro. 6. These data together with lack of tyramine sympathomimetic potentiation by rasagiline, at selective MAO-B inhibitory dosage, indicate that this inhibitor like selegiline may be a useful agent in the treatment of Parkinson's disease in either symptomatic or L-DOPA adjunct therapy, but lack of amphetamine-like metabolites could present a therapeutic advantage for rasagiline.     

Species differences in the deamination of dopamine and other substrates for monoamine oxidase in brain

Cortex and caudate specimens from human, non-human primate and rodent brains were examined for their ability to deaminate dopamine and for their sensitivity to irreversible monoamine oxidase (MAO) inhibitors. Using inhibition curves obtained with clorgyline, deprenyl and pargyline to estimate the relative proportions of MAO-A and MAO-B activity, dopamine was found to be deaminated predominantly by MAO-A in rat cortex and caudate. In contrast, dopamine was primarily an MAO-B substrate in human and vervet cortex and caudate. When clorgyline inhibition curves with tyramine or dopamine as substrate were compared in human, vervet and rat cortex, more pronounced species differences were found with dopamine than with tyramine. In all three species caudate tended to be more sensitive to inhibition by low concentrations of clorgyline than was cortex, suggesting a higher proportion of MAO-A activity in caudate. Similar species differences were also found when MAO-A activities were estimated using serotonin (5-HT): -phenylethylamine (PEA) ratios (5-HT/5-HT + PEA). These ratios with selective substrates were highly correlated with clorgyline inhibition curves obtained with tyramine as substrate across 29 brain regions and tissues from different rodent and primate species (r=0.85, P<0.001). Data from both the substrate ratios and the clorgyline inhibition curves confirmed the relative predominance of MAO-B activity in primate brain regions (70–85%) as compared to rat brain regions (45%). Smaller species differences were observed in liver. Species differences in the proportion of brain MAO-A and B activities and in the deamination of dopamine and other substrates for MAO may have important implications in regard to the widespread use of rodent rather than primate models in the study of biogenic amine metabolism and of drugs affecting amine function.     

Molecular determinants of MAO selectivity in a series of indolylmethylamine derivatives: biological activities, 3D-QSAR/CoMFA analysis, and computational simulation of ligand recognition

A series of indolylmethylamine derivatives were assayed toward MAO-A and MAO-B inhibition. The K(i) values of these compounds are in the range from 0.8 to >10(6) nM for MAO-A or from 0.75 to 476000 nM for MAO-B. The most selective MAO-A or MAO-B inhibitors elicit a ratio of K(i) in the order of 1500 or 1000, respectively. Comparison of MAO-A and MAO-B CoMFA models showed that both the steric and electrostatic properties at the 5 position of the indole ring are determinant for MAO selectivity. Computational simulations of the complex between this part of the ligand and Phe-208 of MAO-A or Ile-199 of MAO-B, experimentally identified as responsible for substrate selectivity, allowed us to further characterize the nature of these enzyme-inhibitor interactions.     

The Anticonvulsant FCE 26743 is a Selective and Short-acting MAO-B Inhibitor Devoid of Inducing Properties towards Cytochrome P450-dependent Testosterone Hydroxylation in Mice and Rats

Abstract— The effects of the potent anticonvulsant FCE 26743 ((S)-2-(4-(3-fluorobenzyloxy)benzylamino)propionamide) on monoamine oxidase (MAO) activity were measured in-vitro and ex-vivo using rat tissue homogenates. In-vitro, FCE 26743 showed potent and selective inhibitory properties towards liver MAO-B, with IC50 values about 10^?7 m for MAO-B and higher than 10^?5 m for MAO-A. When determined ex-vivo in brain, the ED50 value for the inhibition of MAO-B was 1·1 mg kg^?1 (p.o.) 1 h post-dosing, whereas MAO-A remained virtually unaffected after administration of 60 mg kg^?1. Similar effects were seen in liver. Following oral administration of 5 mg kg^?1 FCE 26743 to rats, brain MAO-B inhibition was 79% after 1 h and 13% after 24 h, indicating that FCE 26743 behaves as a short-acting MAO-B inhibitor. The ability of FCE 26743 to act as a MAO substrate was assessed in mice by measuring the urinary excretion of alaninamide, a potential metabolite of FCE 26743 which would result from the action of MAO. No alaninamide was detectable in the 0–8 h urines after administration of a 119 mg kg^?1 dose, suggesting that FCE 26743 is not, or only to a small degree, a substrate of MAO. The effects of FCE 26743 on cytochrome P450 enzymes involved in testosterone hydroxylation were determined in rats after repeated administration. No induction of the cytochrome P450 system was noted.     

Monoamine oxidase in pancreatic islets,exocrine pancreas,and liver from rats. Characterization with clorgyline,deprenyl, pargyline,tranylcypromine, and amezinium

Monoamine oxidase (MAO) was characterized in tissue homogenates from pancreatic islets, exocrine pancreas, and liver from rats. Phenylethylamine was preferentially deaminated by pancreatic islet MAO while 5-hydroxytryptamine was preferentially deaminated by MAO from exocrine pancreas, and tyramine was a good substrate for both tissues. All three substrates were well deaminated by liver tissue. Clorgyline, a selective inhibitor of MAO-A, preferentially inhibited deamination of 5-hydroxytryptamine by all three tissue homogenates, while deprenyl, a selective inhibitor of MAO-B, preferentially inhibited deamination of phenylethylamine. In the case of pargyline, a less selective MAO-B inhibitor, the preference in favour of phenylethylamine was less pronounced. According to these results, MAO in pancreatic islets can be classified as predominantly type B enzyme species and MAO in exocrine pancreas as predominantly type A enzyme species while both types of the enzyme are present in the liver. Using the same three MAO substrates and compared with the effects of the selective enzyme inhibitors, clorgyline and deprenyl, tranylcypromine can be classified as a potent nonselective inhibitor of MAO in homogenates of all three tissues investigated with a slight preference in favour of the inhibition of the B-form of the enzyme, while in contrast amezinium can be classified as a weak nonselective inhibitor of MAO with a slight preference in favour of the inhibition of the A-form of the enzyme. All MAO inhibitors tested also inhibited insulin secretion by isolated incubated rat pancreatic islets, however only at IC50 which were two to three decimal powers higher than those necessary for the inhibition of the MAO activity, thus indicating that inhibition of MAO activity and inhibition of insulin secretion are apparently not closely related.     

Interaction of psychotropic drugs with monoamine oxidase in rat brain

Literature observations indicate that some psychotropic drugs may have inhibitory activity towards monoamine oxidase (MAO). This study was undertaken to assess the potency, isozyme selectivity and mechanism of inhibition of representative first- and second-generation antidepressant drugs towards rat brain MAO-A and MAO-B. Five tricyclic antidepressants (imipramine, trimipramine, clomipramine, amitriptyline and doxepine) and three selective serotonin reuptake inhibitors (fluoxetine, fluvoxamine and citalopram) were examined. They showed inhibitory activity towards MAO-A and MAO-B, with clear selectivity for MAO-B (Ki in the micromolar range). Their mechanism of inhibition was competitive towards MAO-B and of a mixed competitive type towards MAO-A. The results suggest that some of the drugs examined might also contribute an MAO inhibitory effect in chronically treated patients.     

Selectivity of clorgyline and pargyline as inhibitors of monoamine oxidases A and B in vivo in man

During 4 weeks of treatment with clorgyline, a selective MAO-A inhibitor, platelet monoamine oxidase (MAO) activity was unchanged. During a similar 4-week crossover treatment period with pargyline, a selective MAO-B inhibitor, platelet MAO activity was essentially completely inhibited in the same individuals. The differential effects of the two drugs on platelet MAO, which consists exclusively of the MAO-B form, suggests that the in vitro selectivity of clorgyline, and possibly of pargyline, on MAO-A and MAO-B may be maintained in vivo during long-term administration in man. Reductions in blood pressure, heart rate, and plasma amine oxidase activity were generally similar in magnitude during treatment with both drugs, however, suggesting that either these effects are nonspecific consequences of both MAO-A and MAO-B inhibition, or that pargyline also inhibited MAO-A activity.     

Does the B form selective monoamine oxidase inhibitor lose selectivity by long term treatment?

Rats were treated subcutaneously with different doses of the “B form” selective monoamine oxidase (MAO) inhibitors deprenyl (phenylisopropylmethylpropargylamine), U-1424 (N-methyl-N-propargyl-[2-furyl-1-methyl]-ethylammonium) and J-508 (N-methyl-N-propargyl-[1-indenyl]-ammonium. HCl) in order to study the changes of their selectivity during 21 days of treatment. When the daily dose of (?) deprenyl and U-1424 was 0.05 or 0.25 mg/kg body wt (similar to the human dosage of deprenyl in clinical trials), in spite of their repeated administration, a fairly selective inhibition pattern was maintained. In this case at a low rate of oxidation of beta-phenylethylamine (MAO-B form specific substrate) the conversion of serotonin (MAO-A form specific substrate) was near to the untreated value. When 1.0 mg/kg of these inhibitors were repeatedly administered they also inhibited the A form of MAO.As J-508 is a more potent MAO inhibitor than deprenyl and U-1424. even the lowest dose (0.05 mg/kg) used in this study proved to inhibit MAO-A. All the compounds tested were less effective on liver than on brain MAO; thus their selectivity was more pronounced on liver homogenate.     

Effect of diethylnitrosamine on monoamine oxidase in rat liver

The effect of diethylnitrosamine (DEN), a well-known experimental carcinogen, toward MAO-A and MAO-B activity of rat liver was investigated. The oxidations of both beta-PEA (MAO-B) and 5-HT (MAO-A) were inhibited by DEN. The K1 values of DEN in the inhibition of rat liver MAO-A and MAO-B activity were determined. The kinetic data show that DEN is a competitive, MAO-B selective inhibitor and its inhibitory effect on MAO-B is about 4-fold more potent than that on MAO-A. DEN might change the proportions of the multiple forms of MAO activity in tumor cells.     

Tyramine infusions and selective monoamine oxidase inhibitor treatment

The relationship between changes in IV tyramine pressor sensitivity accompanying selective monoamine oxidase (MAO) inhibitor treatment and estimates of MAO-A and MAO-B inhibition in vivo were studied. Reductions in platelet MAO activity provided an index of MAO-B inhibition, while changes in plasma 3-methoxy-4-hydroxyphenethylene glycol (MHPG) were used as an hypothesized reflection of MAO-A inhibition. Chronic treatment with the MAO-A inhibitor clorgyline and the MAO-B inhibitor pargyline showed significant inhibition of the alternate MAO enzyme as well, although this crossover effect was greater for pargyline than clorgyline. The MAO-B inhibitor deprenyl appeared to maintain the greatest degree of MAO inhibition selectivity in vivo. Tyramine pressor sensitivity changes accompanying administration of the MAO inhibitors were highly correlated with decreases in plasma MHPG (r=0.92), supporting our previous data indicating the rank order of clorgyline > pargyline > deprenyl for enhancement of tyramine pressor sensitivity and, thus, suggesting that tyramin potentiation is primarily a function of MAO-A rather than MAO-B inhibition. Changes in plasma MHPG are suggested to provide a potentially useful clinical index of in vivo MAO-A inhibition.Presently with the Biological Psychiatry Branch, NIMH     

Novel monoamine oxidase inhibitors, 3-(2-aminoethoxy)-1,2-benzisoxazole derivatives, and their differential reversibility

Although possible usefulness of non-selective monoamine oxidase (MAO) inhibitors for Parkinson's disease therapy has been suggested in the literature, MAO inhibitors whose inhibition is reversible and have dual action to both MAO-A and -B subtypes is not available yet. Subtype selectivity and reversibility of a series of novel MAO inhibitors, 3-(2-aminoethoxy)-1,2-benzisoxazole derivatives, were studied. Several dual MAO inhibitors, which inhibit both MAO-A and -B, were obtained. When administered to mice, their effects were generally reversible. Among the derivatives, RS-1636 and RS-1653 had much longer duration of brain MAO-B inhibition than that of MAO-A. In vitro, the inhibited MAO-A activity by these compounds was partially recovered by buffer change at 4 degrees C, while little MAO-B activity was recovered. Although it is not fully elucidated yet, the reversibility of these inhibitors is probably determined primarily by this dissociation profile. This unique differential reversibility indicates that optimization of the balance of actions can be achieved by differentiating reversibility to each target molecule.     

Monoamine oxidase A and B inhibitors

This patent update relates to monoamine oxidase inhibitors, covers the patent activity in this field for the last 3 - 5 years and focuses on the most relevant filings. Patents are classified according to the type of MAO inhibition: selective MAO-A and MAO-B inhibitors, and mixed and reversible MAO-A and MAO-B inhibitors.     

次黄嘌呤对单胺氧化酶的抑制作用

实验证明给小鼠po次黄嘌呤25～500 mg/kg时,对肝和脑中单胺氧化酶B(MAOB)活性的抑制作用与剂量成明显的量—效关系,对MAO-A活性的抑制较弱,且无明显的量—效关系。给小鼠一次po次黄嘌呤500 mg/kg,于给药后16h,对MAO抑制作用最明显。sc时,对肝中MAO活性抑制也以给药后16 h最明显,但对脑中MAO活性抑制不明显。离体实验证明,次黄嘌呤对MAO-B的抑制为竞争性,对MAO-A则为混合型抑制。     

Relation between brain monoamine oxidase (MAO) activity and the firing rate of locus coeruleus neurons

Summary Utilizing a specific low substrate concentration technique, intrasynaptosomal MAO-A and MAO-B activities within the rat brain noradrenaline system were studied. It was found that mainly MAO-A was localized intrasynaptosomally, whereas MAO-B contributed with less than 15% of the total intrasynaptosomal MAO activity, a phenomenon that was also observed within the central dopamine system. It is suggested that the intrasynaptosomal pool of MAO in the noradrenaline and the dopamine systems may reflect the density of innervation of the respective system throughout the brain. In addition, the effects of various selective monoamine oxidase (MAO) inhibitors on the noradrenergic intrasynaptosomal MAO activity as well as on the neuronal firing rate of noradrenaline containing cells in the locus coeruleus (LC) were investigated. Pretreatment with the MAO-A selective inhibitors clorgyline (10 mg/kg, i.p., 1 h) or (+)-FLA 336 (1 mg/kg, i.p., 1 h) caused a significant depression (40%) of mean spontaneous firing rate of LC neurones, randomly encountered throughout the LC. The MAO-B selective inhibitor pargyline (10 mg/kg, i.p., 1 h) was found to lack effect in this regard. However, pretreatment with (–)-deprenyl (10 mg/kg, i.p., 1 h), equally a selective MAO-B inhibitor, markedly suppressed the spontaneous firing rate of LC units. This inhibition by (–)-deprenyl was blocked by pretreatment with SK&F 525 A (50 mg/kg, i.p., 30 min), an inhibitor of microsomal drug metabolizing enzymes. Thus, the depression of LC units by (–)-deprenyl seems to be executed by a metabolite, e. g. l-amphetamine. Taken together, the present electrophysiological and biochemical results show that the neuronal depression of noradrenaline neurones in the LC by MAO-inhibitors is specifically related to the inhibition of MAO-A. Furthermore, the data indicate a relationship between the activity of intrasynaptosomally located MAO-A and the neuronal activity of central noradrenaline pathways.     

The evaluation of isatin analogues as inhibitors of monoamine oxidase

The small molecule, isatin, is a well-known reversible inhibitor of the monoamine oxidase (MAO) enzymes with IC₅₀ values of 12.3 and 4.86 μM for MAO-A and MAO-B, respectively. While the interaction of isatin with MAO-B has been characterized, only a few studies have explored structure–activity relationships (SARs) of MAO inhibition by isatin analogues. The current study therefore evaluated a series of 14 isatin analogues as in vitro inhibitors of human MAO-A and MAO-B. The results indicated good potency MAO inhibition for some isatin analogues with five compounds exhibiting IC₅₀ < 1 μM. 4-Chloroisatin ( 1b ) and 5-bromoisatin ( 1f ) were the most potent inhibitors with IC₅₀ values of 0.812 and 0.125 μM for MAO-A and MAO-B, respectively. These compounds were also found to be competitive inhibitors of MAO-A and MAO-B with K_i values of 0.311 and 0.033 μM, respectively. Among the SARs, it was interesting to note that C5-substitution was particularly beneficial for MAO-B inhibition. MAO inhibitors are established drugs for the treatment of neuropsychiatric and neurodegenerative disorders, while potential new roles in prostate cancer and cardiovascular disease are being investigated.     

Styrene inhibits monoamine oxidase A, but not monoamine oxidase B in monkey brain mitochondria

The effects of styrene on mitochondrial monoamine oxidase (MAO) activity in rat and monkey brains were compared in vitro. After preincubation at 25 degrees C for 20 min with 1 mM styrene monomer MAO-A activity in monkey brain was inhibited potently using 5-HT (for MAO-A substrate), but MAO-B activity in monkey brain and platelets were slightly inhibited using beta-PEA (for MAO-B substrate). Styrene monomer also competitively inhibited MAO-A activity in a dose-dependent manner. MAO-A in monkey brain was inhibited by styrene in ascending order of potency: styrene trimer>styrene dimer>styrene monomer. In contrast styrene monomer slightly inhibited both MAO-A and MAO-B activities in rat brain mitochondria. In the present study styrene monomer potently inhibits MAO-A activity, but not MAO-B activity, in monkey brain mitochondria in vitro. These results indicate the inhibiting action of styrene differs depending on animal species and MAO isoforms.     

Stereoselectivity and isoenzyme selectivity of monoamine oxidase inhibitors. Enantiomers of amphetamine, N-methylamphetamine and deprenyl

The enantiomers of amphetamine, N-methylamphetamine and deprenyl were studied, using a solubilised rat liver mitochondrial monoamine oxidase (MAO) preparation, as competitive inhibitors of MAO-A and MAO-B (5-hydroxytryptamine and beta-phenylethylamine as substrate respectively). Only in the case of deprenyl enantiomers inhibiting MAO-B was a preference shown towards the [R]-configuration enantiomer justifying the use of [R]-(-)-deprenyl (as compared to the racemate) for the specific inhibition of MAO-B. Recalculation of the observed Ki values in terms of the base form of the inhibitor indicated that the activity of all enantiomers fell within a narrow, approximately 25-fold range when inhibiting MAO-B. The selectivity of inhibition of MAO-B by [R]-(-)-deprenyl cannot therefore be attributed to any specific structural features of the MAO-B isoenzyme form but rather to a lack of affinity of this enantiomer towards MAO-A.