Distribution of the Two Forms of Monoamine Oxidase Within Monoaminergic Neurons of the Guinea Pig Brain

The relative distribution of type A and type B monoamine oxidase (MAO) inside and outside the monoaminergic synaptosomes in preparations from hypothalamus and striatum of the guinea pig was determined by incubation of synaptosomal preparations of these regions with low concentrations of [14C]5-hydroxytryptamine (5-HT), noradrenaline, and dopamine. The deamination within the monoaminergic synaptosomes was hindered by selective amine uptake inhibitors. In the absence of these inhibitors, both intra- and extraneuronal deamination was measured. The two forms of the enzyme were differentiated with the irreversible and selective MAO-A and MAO-B inhibitors clorgyline and selegiline (l-deprenyl), respectively. [14C]5-HT was deaminated greater than 90% by MAO-A both inside and outside the 5-hydroxytryptaminergic synaptosomes prepared from the guinea pig hypothalamus. The deamination of [14C]noradrenaline within the noradrenergic synaptosomes of the hypothalamic preparation was in the ratio 75:25% for MAO-A:MAO-B; the corresponding ratio outside these synaptosomes was 45:55%. The deamination of [14C]dopamine within dopaminergic synaptosomes in the striatal preparation was 65% type A:35% type B, whereas outside these synaptosomes the ratio was 35:65%. Because the relative amounts and the distribution of the two forms of MAO in the guinea pig brain seem to be similar to those previously detected for the human brain, the MAO in the guinea pig brain may be a good model for the MAO in the human brain.     

Changes in brain monoamine oxidase activity in conditioned passive avoidance reaction in rats]

Activity of the enzyme monoamine oxidase (MAO) and kinetic parameters (Km, Vmax) for the 5-hydroxytryptamine (5-HT) and dopamine deamination were examined in the brain of rats with conditioned passive avoidance recall. Changes of the 5-HT and dopamine deamination were found in amygdala, striatum and frontal cortex. MAO activity was not changed in hippocampus. In amygdala the rate of 5-HT deamination was significantly increased and kinetic studies revealed increased affinity of the enzyme for 5-HT. The metabolism of dopamine in amygdala was unchanged. In frontal cortex the deamination of 5-HT was not changed, but the dopamine deamination significantly decreased. This decrease was due to lowering of MAO affinity for dopamine. In striatum the metabolism of both 5-HT and dopamine was reduced, and kinetic studies showed the lowering of Vmax for 5-HT and dopamine deamination.     

Change in serotonin metabolism in the rat brain in response to the repeated stimulus presentation

The content of serotonin (5-HT), its metabolite 5-hydroxyindoleacetic acid (5-HIAA), monoamine oxidase (MAO) activity and kinetic parameters (K(m) and Vmax) for the reaction of 5-HT deamination, were examined in various regions of the rat brain after repeated presentation of a contextual stimulus. Habituation to the stimulus was accompanied by an increase of 5-HT metabolism and active transport of 5-HIAA in the amygdala, striatum and midbrain, while these changes were not found in the prefrontal cortex and hippocampus. Kinetic studies have revealed that the enhancement of 5-HT deamination by MAO in the brain structures was mediated by different catalytic mechanisms. A significant decrease in K(m) value for 5-HT deamination in the amygdala indicated an increase in the affinity of enzyme towards 5-HT. In the striatum the enhanced MAO activity was provided by increasing maximal rate of 5-HT deamination. It is concluded that an activation of presynaptic mechanisms of the serotonergic transmission in the amygdala and striatum is involved in the inhibition of biological significance and attention to repeated presentation of stimulus.     

Reaction of deamination of monoamines in the brain and heart of rats under the toxic action of hyperbaric oxygenation

Kinetic parameters of monoamine deamination processes in the rat brain and heart after hyperbaric oxygenation (HBO) in toxic conditions (6 ata) were studied. HBO was shown to cause a substantial reduction in MAO affinity to serotonin in the brain, but not in the heart. Contrastingly, MAO affinity to dopamine was found to decrease in the heart, but not in the brain in response to HBO. Differences of tyramine and 2-phenylethylamine deamination in the rat brain and heart were also reciprocal following toxic HBO. In the initial phase of seizure episode MAO activity in the brain and heart was also different. Distinct mechanisms of adaptation to toxic oxygen in the central nervous system and cardiovascular system are discussed.     

Kinetics of the oxidative deamination reaction in the preconvulsive period of oxygen-induced epilepsy

Kinetic parameters of monoamine oxidative deamination in compensatory and preconvulsive periods of oxygen epilepsia were studied. It was shown that in rat brain MAO's affinity for serotonin reduced from the 5th minute of exposure to hyperbaric oxygen and went on reducing on the 15th minute. In rat heart the affinity of MAO for serotonin firstly decreased and then returned to normal meaning. Dopamine deamination in rat brain in compensatory period of epilepsia was activated and then was inhibited. In rat heart from the 5th minute of exposure to oxygen dopamine and 2-phenylethylamine deamination was blocked. Tyramine deamination in preconvulsive period of epilepsia changed in a complex manner. It is concluded that the kinetic parameters of monoamine deamination change in the initial phases of exposure to hyperbaric oxygen, and the most distinct modifications take place in rat heart, but not in rat brain.     

A and B Forms of Monoamine Oxidase Within the Monoaminergic Neurons of the Rat Brain

The inhibition of the A and B forms of monoamine oxidase (MAO) inside and outside serotonergic, noradrenergic, and dopaminergic synaptosomes in homogenates of rat hypothalamus or striatum by clorgyline, a selective and irreversible MAO-A inhibitor, and selegiline, a selective and irreversible MAO-B inhibitor, was examined. Intrasynaptosomal deamination at low concentrations of the substrates [14C]5-hydroxytryptamine ([14C]5-HT; 0.1 microM), [14C]noradrenaline (0.25 microM), [14C]3,4-dihydroxyphenylethylamine ([14C]dopamine; 0.25 microM), and [14C]tyramine (0.25 microM) was hindered by selective uptake inhibitors (citalopram, maprotiline, and amfonelic acid) in the incubation media. Thus, the difference between the deamination of 14C-amine in the absence and presence of the appropriate selective uptake inhibitor provided a measure of deamination in the specific aminergic synaptosomes. This was verified by determining the loss of MAO activity within noradrenergic and serotonergic systems after degeneration of the nerve terminals by the neurotoxins N-chloroethyl-N-ethyl-2-bromobenzylamine and p-chloroamphetamine. Results with the two inhibitors revealed that the A and B forms were responsible for 80 and 20%, respectively, of the deamination of [14C]5-HT within serotonergic synaptosomes from the hypothalamus. The deamination of [14C]noradrenaline within the noradrenergic synaptosomes from the hypothalamus and that of [14C]dopamine and [14C]tyramine within the striatal dopaminergic synaptosomes were due to MAO-A. About 10% of the deamination of [14C]noradrenaline, [14C]dopamine, and [14C]tyramine outside the noradrenergic or dopaminergic synaptosomes was brought about by the B form, with the remainder being deaminated by MAO-A.     

Guinea Pig Striatum as a Model of Human Dopamine Deamination: The Role of Monoamine Oxidase Isozyme Ratio, Localization, and Affinity for Substrate in Synaptic Dopamine Metabolism

The kinetic properties of type A and type B monoamine oxidase (MAO) were examined in guinea pig striatum, rat striatum, and autopsied human caudate nucleus using 3,4-dihydroxyphenylethylamine (dopamine, DA) as the substrate. MAO isozyme ratio in guinea pig striatum (28% type A/72% type B) was similar to that in human caudate nucleus (25% type A/75% type B) but different from that in rat striatum (76% type A/24% type B). Additional similarities between guinea pig striatum and human caudate nucleus were demonstrated for the affinity constants (Km) of each MAO) isozyme toward DA. Endogenous concentrations of DA, 3-methoxytyramine, 3,4-dihydroxyphenylacetic acid, and homovanillic acid were also measured in guinea pig and rat striatum following selective type A (clorgyline-treated) and type B (deprenyl-treated) MAO inhibition. In guinea pig, DA metabolism was equally but only partially affected by clorgyline or deprenyl alone. Combined treatment with clorgyline and deprenyl was required for maximal alterations in DA metabolism. By contrast, DA metabolism in rat striatum was extensively altered by clorgyline but unaffected by deprenyl alone. Finally, the deamination of DA in synaptosomes from guinea pig striatum was examined following selective MAO isozyme inhibition. Neither clorgyline nor deprenyl alone reduced synaptosomal DA deamination. However, clorgyline and deprenyl together reduced DA deamination by 94%. These results suggest that the isozyme localization and/or isozyme affinity for DA, rather than the absolute isozyme content, determines the relative importance of type A and type B MAO in synaptic DA deamination. Moreover, based on the enzyme kinetic properties of each MAO isozyme, guinea pig striatum may serve as a suitable model of human DA deamination.     

Distinctions in metabolism of serotonin and dopamine in rat brain during latent inhibition

Latent inhibition (LI) is a behavioral phenomenon, in which repeated presenting of a non-reinforced stimulus retards conditioning to this stimulus when it is coupled with a reinforcer. In order to find specific serotonin (5-HT- and dopamine (DA) changes mediating the LI, the 5-HT and DA metabolism was investigated in certain brain regions. Oxidative deamination of 5-HT and DA by monoamine oxidase (MAO) was determined in the prefrontal cortex, striatim, amygdala, and hippocampus at preexposure and testing stages of the LI using the passive avoidance procedure in rats. Preexposed animals demonstrated high MAO activity for 5-HT deamination in the amygdala and striatum and lower MAO activity for DA deamination in the amygdala and hippocampus. After testing the LI, a high level of 5-HT deamination by MAO was revealed in the amygdala, white the lower level of 5-HT deamination by MAO was shown in the prefrontal cortex. At the same time, no changes in DA metabolism were found in all the brain regions studied. Thus, the role of dopaminergic system in the LI effect may be limited by the preexposure stage. The obtained evidence suggests that the enhanced 5-HT activity in the amygdala and striatum induced by the preexposed stimulus is a principal biochemical mechanism underlying the LI.     

THE EFFECTS OF CHRONIC REGIMENS OF CLORGYLINE AND PARGYLINE ON MONOAMINE METABOLISM IN THE RAT BRAIN

The effects of chronic administration of clorgyline and pargyline on rat brain monoamine metabolism have been examined. The inhibitory selectivity of these drugs towards serotonin deamina-tion (MAO type A) and phenylethylamine deamination (MAO type B) can be maintained over a 21-day period by proper selection of low doses of these drugs (0.5-1.0 mg/kg/24h). The results are consistent with MAO type A catalyzing the deamination of serotonin and norepinephrine and with MAO type B having little effect on these monoamines. Dopamine appears to be dcaminated in vivo principally by MAO type A. Clorgyline administration during a 3-week period was accompanied by persistent elevations in brain norepinephrine concentrations; serotonin levels were also increased during the first 2 weeks, but returned towards control levels by the third week of treatment. Low doses of pargyline did not increase brain monoamine concentrations, but treatment with higher doses for 3 weeks led to elevations in brain norepinephrine and 5-hydroxytryptamine; at this time significant MAO-A inhibition had developed. The changes in monoamine metabolism seen at the end of the chronic clorgyline regimen are not due to alterations in tryptophan hydroxylase activity. At this time tyrosine hydroxylase activity was also unaffected.     

Effect of prolonged cold exposure on monoamine oxidase activity and kinetics and on serotonin metabolism in the rat brain

Effects of long-term cold exposure on the content of serotonin and its metabolite 5-hydroxyindolacetic acid (5-HIAA) and monoamine oxidase (MAO) activity and kinetic parameters (Km and Vmax) of oxidative deamination of serotonin in rat brain stem. The increase of 5-HIAA level in the initial period of chronic cold exposure was determined by the blockade of active metabolite transport from the brain. The level of serotonin and the rate of its catalytic deamination by MAO were found to be decreased in cold-adapted rats. The magnitude of the Km of serotonin deamination was unchanged.     

Effects of intrastriatal kainic acid injection on [3H]dopamine metabolism in rat striatal slices: evidence for postsynaptic glial cell metabolism by both the type A and B forms of monoamine oxidase

Abstract: Intrastriatal injections of kainic acid (KA) were utilized to investigate the cellular localization of postsynaptic dopamine (DA) metabolism by type A and B monoamine oxidase (MAO) in rat striatum. At 2 days postinjection, maximal degeneration of cholinergic and γ-aminobutyric acid (GABA)ergic neurons was observed and found to be associated with a significant decrease in both type A and B MAO activity. However, over the next 8-day period, when only the process of gliosis appeared to be occurring, a selective return to control of type B MAO activity was seen. When the metabolism of [³H]DA (10^?7 M) was examined in 8-day KA-lesioned rat striatal slices, an increase in [³H]dihydroxyphenylacetic acid (DOPAC) and [³H]homovanillic acid (HVA) formation was observed. The KA-induced elevation of [³H]DOPAC formation (but not [³H]HVA) was abolished by the DA neuronal uptake inhibitor nomifensine. This is consistent with earlier findings suggesting that HVA is formed exclusively within sites external to DA neurons. Experiments with clorgyline and/or deprenyl revealed that the relative roles of type A and B MAO in striatal DA deamination remained unchanged following KA (90% deamination by type A MAO) even though total deamination was substantially enhanced. At high concentrations of [³H]DA (10^?5 M), deamination by type B MAO could be increased to 30% of the total MAO activity; however, this was observed in both control and KA-lesioned striata. These results suggest that KA-sensitive neurons contain type A and/or type B MAO. Moreover, whereas these neurons may metabolize DA, a major portion of postsynaptic DA deamination appears to occur within glial sites of rat striatal tissue. Furthermore, glial cells would appear to contain functionally important quantities of both type A and B MAO.     

THE METABOLISM OF TRITIATED DOPAMINE IN REGIONS OF THE RAT BRAIN IN VIVO—II

Abstract— The levels of tritiated catecholamines and metabolites were measured in regions of the rat brain at intervals after the intraventricular injection of [³H]dopamine, [³H]nor-adrenaline and [³H]normetanephrine. The disappearance of catecholamines and appearance of metabolites with time and the regional turnover rates of these amines indicate that the major pathway of the metabolism of noradrenaline and dopamine actively released from physiological storage sites is to the neutral alcoholic metabolites. The acid metabolites, homovanillic acid and 3,4-dihydroxyphenylacetic acid appear to be only minor products of normal dopamine metabolism in rat brain regions including the striate, but are the main end products of the metabolism of excess exogenous dopamine.
The active metabolism of stored noradrenaline to alcohol metabolites is also indicated by the increase in neutral alcohol metabolites accompanying the increased noradrenaline turnover when rats were subjected to electroshock stress. Therefore in the rat brain, neutral alcohol metabolites of dopamine and noradrenaline have great significance in the study of physiological catecholamine turnover in any region.     

The oxidation of adrenaline and noradrenaline by the two forms of monoamine oxidase from human and rat brain

The selective monoamine oxidase inhibitors clorgyline and (−)-deprenyl were used to study the distribution of monoamine oxidase-A and -B (MAO-A, MAO-B) activities towards (−)-noradrenaline and (+),(−)-adrenaline in homogenates from seven different regions of human brain. The activities towards 5-hydroxytryptamine and 2-phenethylamine, which are essentially specific substrates for the A- and B-forms, respectively, under the conditions used in this work, were also determined. Noradreanline and adrenaline were substrates for both forms of the enzyme in all regions studied. The total MAO activity was found to be highest in the hypothalamus and lowest in the cerebellar cortex. Use of the selective MAO inhibitors clorgyline and (−)-deprenyl also showed adrenaline and noradrenaline to be substrates for both forms of the enzyme in rat brain. In human cerebral cortex and rat brain the two forms were found to have similar K_m-values and maximum velocities towards adrenaline. These values for the two forms were also found to be similar in human cerebral cortex when noradrenaline was used as the substrate. In contrast MAO-A showed a significantly lower K_m and a higher maximum velocity towards noradrenaline in rat brain. These results suggest that the rat may not provide a close model of the human for studies on the effects of MAO inhibitors on brain noradrenaline metabolism.     

Monoamine oxidase in adult Ascaridia galli

Monoamine oxidase (MAO), catalysing oxidative deamination of biogenic monoamines, has been detected in adult Ascaridia galli. MAO was present in mitochondria and deaminated noradrenaline at the maximal rate, although serotonin, adrenaline, tyramine and dopamine were also degraded but more slowly. Of the organs studied, the body wall, female reproductive organ and intestine, the body wall (containing neuronal structures) showed highest MAO activity. Km value for chick ascarid mitochondrial MAO using tyramine as substrate was 1.66 X 10(-3) M and it was most active at 2.5 mM tyramine concentration, pH 7.5 and 40 degrees C. MAO of A. galli appeared to be thermolabile as nearly 80% of its activity was lost when the incubation temperature was increased 5 degrees above optimum.     

Inhibition of brain monoamine oxidase in the rat by multiple pyrazidol administration

Pyrazidol, which is chemically 2,3, 3a, 4, 5, 6-hexahydro-8-methyl-1H-pyrazino[3,2,1-j,k] carbazole hydrochloride (international name pirlindole) administered repeatedly (21 days) to rats at a dose of 25 mg/kg per os maintained the selectivity of its inhibitory effect toward type A MAO. When administered repeatedly the inhibitory effect of pyrazidol was 1.5-2-fold higher than after a single administration. The effect of pyrazidol on rat brain MAO was reversible whatever the route of administration. The enzymatic activity returned to normal within 24 h after the last administration. The data obtained suggest that the capacity of selective inhibiting the deamination of the neurotransmitters such as serotonin and noradrenaline in human brain is of paramount importance for therapeutic effect of pyrazidol.     

The action of befol and its derivatives on monoamine oxidase of different origins

The effect on deamination of serotonine, dopamine, tiramine and 2-phenylamine of benzamide derivatives befol, moclobemide and LIS-641 was studied. Befol and moclobemide are inhibitors of serotonine deaminating activity of MAO. The different sensitivity of this activity to the effect of the benzamide derivatives in beef or rat brain and human placenta was noted. The inhibition was more distinct in tissue homogenate than in corresponding mitochondrial fractions.     

Inhibition of Monoamine Oxidase by 7-Chloro-4-Nitrobenzofurazan

7-Chloro-4-nitrobenzofurazan (NBD-Cl) is a potent inhibitor of both types of monoamine oxidase (MAO). NBD-Cl competitively inhibited the oxidative deamination of kynuramine catalyzed by human placenta MAO-A, the oxidative deamination of benzylamine catalyzed by bovine liver MAO-B, the oxidative deamination of serotonin catalyzed by rat brain MAO-A, and the oxidative deamination of phenylethylamine catalyzed by rat brain MAO-B. In addition, a time-dependent inactivation of MAOs by NBD-Cl has been demonstrated upon incubation of the enzyme preparations with NBD-Cl at pH 9, but not at pH 7.5. The time-dependent inhibition of MAO by NBD-Cl could be prevented by the addition of 4-nitrophenyl azide, an active site-directed label of MAO, during incubation of the enzyme with NBD-Cl. On the basis of these findings, it is suggested that at pH 9, NBD-Cl modifies one (or more) essential lysine residue(s) in the active sites of the two types of MAO.     

Relationship Between Catechol-O-Methyltransferase and Phenolsulfotransferase in the Metabolism of Dopamine in the Rat Brain

The relationship between phenolsulfotransferase (PST) and catechol-O-methyltransferase (COMT) in the metabolism of free 3,4-dihydroxyphenylethylamine (DA, dopamine) in the rat brain was studied. In rats not pretreated with a monoamine oxidase (MAO) inhibitor a huge increase of free DA in the brain, following an intraperitoneal injection of L-3,4-dihydroxyphenylalanine (L-DOPA) or an intraventricular injection of free DA, did not lead to any noticeable change in DA sulfate or 3-methoxytyramine (3-MT), which remained undetectable by the present HPLC method. However, in rats previously treated with the MAO inhibitors pargyline or tranylcypromine, the same L-DOPA or free DA treatment resulted in significant increases in both 3-MT and DA sulfate in the hypothalamus, brainstem, and striatum. This response of COMT and PST was not affected by prior treatment of the rats with 6-hydroxydopamine, which suggests that O-methylation and sulfoconjugation occur outside adrenergic neurons not destroyed by the neurotoxin. Inhibition of COMT activity did not lead to any increase in DA sulfate, which showed that despite their common mode of action (both enzymes react preferentially at the same hydroxyl group in the DA molecule), the two enzymes are not competitive. After MAO inhibition there were strong correlations between an increase in DA sulfate and 3-MT on the one hand, and between free DA and 3-MT on the other. Because 3-MT is a marker of central DA release, these data suggest that inhibition of MAO activity not only affects DA metabolism by this enzyme but also influences DA release in the rat brain.     

THE EFFECT OF HYPOXIA ON MONOAMINE SYNTHESIS, LEVELS AND METABOLISM IN RAT BRAIN

Abstract— Rats were exposed to 5.6% oxygen environments for up to 2 h. The accumulation of brain DOPA and 5-hydroxytryptophan at 30 min after decarboxylase inhibition was used to estimate cerebral tryosine and tryptophan hydroxylase activity, respectively, in vivo. There was a continuing decrease in tryosine hydroxylase activity during the 2 h in whole brain as well as five brain regions. Tryptophan hydroxylase activity declined during the 1st h, but then increased towards control levels during the 2nd h. There was an increase in brain tryptophan during the 2nd h as well. In whole brain and the five brain regions, there was no significant change in the levels of noradrenaline, dopamine or 5-hydroxytrypamine. During a 1 h exposure to 5.6% oxygen, there was decreased accumulation of noradrenaline, dopamine and 5-hydroxytryptamine after MAO inhibition and decreased accumulation of homovanillic acid and 5-hydroxyindoleacetic acid after probenecid administration. The dercreased synthesis and metabolism of the monoamines is most likely attributable to insufficient brain tissue oxygen as a substrate for the two hydroxylase enzymes.     

Metabolism of octopamine in vitro by monoamine oxidase in some rat tissues.

The deamination $\text{in vitro}$ of DL-octopamine by MAO in rat brain, heart, kidney, liver and vas deferens has been studied by a radiochemical method. Kinetic constants for octopamine metabolism, as well as its sensitivity to inhibition by the irreversible MAO inhibitor clorgyline are described for each tissue. On the basis of the inhibition data, it was concluded that octopamine is metabolized preferentially by type A MAO in heart, kidney and vas deferens. However, in brain and liver, type B MAO is also responsible for a significant proportion of total octopamine metabolism. These studies are discussed in relation to current ideas about the regulation of octopamine concentrations in animal tissues, and the possible importance of this amine in mammalian physiology.