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PubMed2004

Dual functions of transcription factors, transforming growth factor-beta-inducible early gene (TIEG)2 and Sp3, are mediated by CACCC element and Sp1 sites of human monoamine oxidase (MAO) B gene.

Monoamine oxidases (MAO) A and B catalyze the oxidative deamination of many biogenic and dietary amines. Abnormal expression of MAO has been implicated in several psychiatric and neurodegenerative disorders. Human MAO B core promoter (-246 to -99 region) consists of CACCC element flanked by two clusters of overlapping Sp1 sites. Here, we show that cotransfection with transforming growth factor (TGF)-beta-inducible early gene (TIEG)2 increased MAO B gene expression at promoter, mRNA, protein, and catalytic activity levels in both SH-SY5Y and HepG2 cells. Mutation of the CACCC element increased the MAO B promoter activity, and cotransfection with TIEG2 further increased the promoter activity, suggesting that CACCC was a repressor element. This increase was reduced when the proximal Sp1 overlapping sites was mutated. Similar interactions were found with Sp3. These results showed that TIEG2 and Sp3 were repressors at the CACCC element but were activators at proximal Sp1 overlapping sites of MAO B. Gel-shift and chromatin immunoprecipitation assays showed that TIEG2 and Sp3 bound directly to CACCC element and the proximal Sp1 sites in both synthetic oligonucleotides and natural MAO B core promoter. TIEG2 had a higher affinity to Sp1 sites than CACCC element, whereas Sp3 had an equal affinity to both elements. Thus, TIEG2 was an activator, but Sp3 had no effect on MAO B gene expression. This study provides new insights into MAO B gene expression and illustrates the complexity of gene regulation.

Apoptosis Regulatory Proteins↗

PubMed2001

Molecular characterization of monoamine oxidases A and B.

Monoamine oxidase A and B (MAO A and B) are the major neurotransmitter-degrading enzymes in the central nervous system and in peripheral tissues. MAO A and B cDNAs from human, rat, and bovine species have been cloned and their deduced amino acid sequences compared. Comparison of A and B forms of the enzyme shows approximately 70% sequence identity, whereas comparison of the A or B forms across species reveals a higher sequence identity of 87%. Within these sequences, several functional regions have been identified that contain crucial amino acid residues participating in flavin adenine dinucleotide (FAD) or substrate binding. These include a dinucleotide-binding site, a second FAD-binding site, a fingerprint site, the FAD covalent-binding site, an active site, and the membrane-anchoring site. The specific residues that play a role in FAD or substrate binding were identified by comparing sequences in wild-type and variants of MAO with those in soluble flavoproteins of known structures. The genes that encode MAO A and B are closely aligned on the X chromosome (Xp11.23), and have identical exon-intron organization. Immunocytochemical localization studies of MAO A and B in primate brain showed distribution in distinct neurons with diverse physiological functions. A defective MAO A gene has been reported to associate with abnormal aggressive behavior. A deleterious role played by MAO B is the activation of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a proneurotoxin that can cause a parkinsonian syndrome in mammals. Deprenyl, an inhibitor of MAO B, has been used for the treatment of early-stage Parkinson's disease and provides protection of neurons from age-related decay.

Amino Acid Sequence↗

PubMed2004

The effect of diabetes mellitus on the morphology and physiology of monoamine oxidase in the pancreas.

Monoamine oxidase (MAO) is an ubiquitous, non-soluble, membrane-bound enzyme, located in the outer membrane of mitochondria. MAO consists of two subtypes, MAO-A and MAO-B, depending on their substrates and sensitivity to inhibitors. MAO consists of two units joined together by a disulphide bond. The two units of MAO and flavin adenine dinucleotide (FAD) form a polymer in the outer membrane of mitochondria. The function of MAO-A is highly dependent on the lipid constituent of mitochondrial membrane, whereas the function of MAO-B does not depend on the lipid status of mitochondrial membrane. Hydrogen peroxide and ammonia are generated during MAO-induced metabolism of its substrates. MAO and its substrates are present in both the exocrine as well as the endocrine parts of the pancreas. In the islet of Langerhans, MAO-A is observed in about 50% of the cells, whereas MAO-B is less abundant and located mainly in the periphery of pancreatic islets. MAO-B is also demonstrated in centroacinar cells and in pancreatic ducts. Electron microscopy studies suggest that MAO is co-localised with insulin in secretory granules of pancreatic beta cells. Pharmacologically, beta-2-adrenoreceptors agonists such as terbutaline can stimulate MAO activity. In contrast, cholinergic muscarinic stimulation does not affect islet MAO activity. MAO activity in pancreatic tissue is significantly reduced in diabetes. This decrease in MAO activity is associated with an increase in pancreatic tissue levels of adrenaline (ADR) and noradrenaline (NA). Studies on the level of 5-hydroxyindoleacetic acid of pancreatic tissues suggest that serotonin level is also increased in diabetics. Many studies show that MAO inhibits insulin secretion. However, some of its substrates including, serotonin, adrenaline and noradrenaline have been shown to stimulate insulin secretion. In conclusion, the activity and subcellular localisation of MAO suggests that MAO may play an important role in pancreatic beta cell function and hence in the pathogenesis of diabetes mellitus.

Animals↗

PubMed1975

Procaine HCl (Gerovital H3): a weak, reversible, fully competitive inhibitor of monoamine oxidase.

A specially stabilized form of procaine hydrochloride (Gerovital H3) has been shown to be a more potent inhibitor of monoamine oxidase than procaine HCl itself and a weaker inhibitor of this enzyme than iproniazid. This preparation was studied to determine its mode of interaction with monoamine oxidase using purified rat brain mitochondrial monoamine oxidase as the enzyme source. Reaction velocities were determined spectrophotometrically by quantitating the rate of appearance of 4-hydroxyquinoline from kynuramine. Dilutional studies comparing the mechanism of inhibition of monoamine oxidase produced by Gerovital H3 and by ipronizid demonstrated that Gerovital H3 was a reversible inhibitor of monoamine oxidase. Analysis of studies using Lineweaver-Burk and Dixon plots revealed that Gerovital H3 was a fully competitive inhibitor of monoamine oxidase. That Gerovital H3 is a weak, reversible, competitive inhibitor of monoamine oxidase may explain the absence of adverse reactions associated with the clinical use of Gerovital H3 as compared to the severe adverse reactions that have been associated with the use of irreversible monoamine oxidase inhibitors.

Animals↗

PubMed1984

Comparison of the properties of semipurified mitochondrial and cytosolic monoamine oxidases from rat brain.

Mitochondrial and cytosolic monoamine oxidases were purified 220- and 129-fold, respectively, from rat brain. The purification procedure involved extraction (without the use of detergents for mitochondrial monoamine oxidase), ammonium sulfate precipitation, and chromatography on Sephadex G-25 and a DEAE-cellulose column. The properties of both enzymes with kynuramine as substrate, including Km values and pH optima at different kynuramine concentrations; the Rf values on polyacrylamide gel electrophoresis; and the thermal inactivation patterns were different. 2-Mercaptoethanol, together with heat treatment, released the flavin and decreased the enzyme activity differentially for the two enzymes. The absorption spectrum showed a "Red shift" in the absorption maxima when the spectra of the non-Triton-treated purified preparations were compared with those of the Triton-treated ones, thus possibly revealing that the mitochondrial and the cytosolic monoamine oxidases may be two different enzyme entities.

Animals↗

PubMed1986

[Formation of the superoxide anion radical during the oxidation of biogenic amines catalyzed by mitochondrial monoamine oxidase].

The studies on the activity of monoamine oxidase from human placenta, using 2-phenylethylamine as a substrate, corroborate the hypothesis on the possible superoxide radical generation upon FAD oxidation at the second (aerobic) stage of monoamine oxidase reaction. It has been shown that hydrogen peroxide, but not other activated O2 forms, was the end product of this reaction. No superoxide radical generation took place in such systems. And therefore, the induction of lipid peroxidation in the presence of catalase was impossible in mitochondrial membranes containing monoamine oxidase and amines oxidized by it.

Biogenic Amines↗

PubMed1994

Cytochemical localization of monoamine oxidase in the pig pineal gland.

Monoamine oxidase plays an important role in the regulation of monoamine levels in the pineal gland by oxidative deamination of serotonin to 5-hydroxyindole acetic acid. The ultrastructural sites of this process are still not well known. In the present study the pig pineal glands were examined by using two cytochemical methods for demonstration of MAO activity at the ultrastructural level. The use of selective inhibitors of MAO-A (clorgyline and amiflamine) and MAO-B (deprenyl) showed that MAO-A was localized predominantly in noradrenergic nerve terminals as well as in the cell membrane of endothelial cells. MAO-B activity was localized in pinealocytes. However, in the outer membranes of cytoplasmic dense bodies, which are characteristic cytoplasmic structures of pig pinealocyte, we noticed the activity of both MAO-A and MAO-B enzymes. The MAO-A was localized exclusively in dense bodies within bulbous ending of pinealocyte processes present in the perivascular space. Our results show that in pig pineal gland there are two forms of MAO localized in different compartments of the pineal tissue. The most important result of our study is the presence of MAO in cytoplasmic dense bodies, which points to these bodies as to the sites of oxidative deamination and confirms their participation in the secretory process in pig pineal gland.

Animals↗

PubMed1991

Distribution of type B monoamine oxidase immunoreactivity in the cat brain with reference to enzyme histochemistry.

We studied the detailed distributions and morphology of structures immunoreactive to type B monoamine oxidase, and compared them with those stained by monoamine oxidase enzyme histochemistry in the brain of cats treated with or without colchicine. By means of the indirect immunohistochemical method in conjunction with type B monoamine oxidase monoclonal antibody, we demonstrated type B monoamine oxidase immunoreactivity in neuronal cell bodies, fibers and astroglial cells in the cat brain. As expected, the distribution of type B monoamine oxidase-immunoreactive cell bodies overlapped that of serotonin-containing ones in the lower brainstem and midbrain, as well as that of histaminergic ones in the posterior hypothalamus. We found novel cell groups containing type B monoamine oxidase in the areas described below. Intense type B monoamine oxidase-immunopositive and enzymatically active neurons, corresponding to liquor-contact ones, were discovered in the wall of the central canal of the spinomedullary junction. Weak immunoreactivity was identified in neurons of the dorsal motor nucleus of the vagus, parvocellular reticular formation and locus coeruleus complex, which have been reported to contain type A monoamine oxidase enzymatic activity. Type B monoamine oxidase-immunostaining in these structures was enhanced by treatment with colchicine. In addition, lightly immunostained cells were distinguished in the caudal portion of the hypothalamic arcuate nucleus, area of tuber cinereum, retrochiasmatic area, and rostral portion of the paraventricular thalamic nucleus after colchicine treatment. These cells also displayed monoamine oxidase activity; however, it was difficult to enzymatically characterize their nature due to its weak activity and sensitivity to inhibitors of both A and B. Distinct type B monoamine oxidase-immunoreactive fibers and terminal-like dots were abundant in the whole brain, particularly in the central gray, dorsal pontine tegmentum, interpeduncular and pontine nuclei, nucleus of the solitary tract and dorsal motor nucleus of vagus, where dense innervations of serotonergic fibers have been reported. Their immunoreactive density increased after colchicine treatment, but monoamine oxidase enzymatic reaction did not. An intense immunoreactivity could be seen in many glial cells in parts of the brain including myelinated axon pathways. The densest accumulation of such labeled glial cells was found in the central gray, inferior olive, medial geniculate body, substantia nigra, ventral tegmental area of Tsai, retrorubral area, hypothalamus, thalamus and bed nucleus of the stria terminalis. In contrast, the striatum contained less numerous type B monoamine oxidase-immunoreactive and enzymatically active astroglial cells in comparison with the other structures.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1987

Fate of the monoamine oxidase inhibitor pargyline in cultured hepatocytes.

The interaction of the monoamine oxidase inhibitor pargyline with cultured hepatocytes has been studied. [Phenyl-3, benzyl-3H] pargyline (38 nM) rapidly enters the cells and a plateau of incorporation into a trichloroacetic acid insoluble form (monoamine oxidase) is reached after 2 hr. The level of labelling is lower in freshly isolated cells than in those in later culture. The maximum incorporation accounts for only 6% of the added radioactivity and produces a 9% inhibition of monoamine oxidase activity. The remaining [3H] pargyline is metabolized and quickly accumulates in the cell culture medium in a form which cannot label exogenous mitochondria. The metabolism of pargyline varies both qualitatively and quantitatively with culture age. In 0 hr and 20 hr-cultured cells one metabolite preferentially appears whilst in 140 hr cultured hepatocytes at least three metabolites are formed. The metabolism of [3H] pargyline in early culture is consistent with a cytochrome P-450 involvement. The use of [3H] pargyline to label monoamine oxidase in cultured hepatocytes offers several attractive features for studying the turnover of this enzyme. These include speed of interaction, non-reutilization, application to normal cells, controlled inhibition of monoamine oxidase and metabolism of non-specific label.

Animals↗

PubMedDate not supplied

[Isolation, purification and the physicochemical properties of the monoamine oxidase from the myocardium].

A preparation of monoamine oxidase, purified 20-fold with a yield of 20%, was isolated from mitochondrial fraction of bovine heart after treatment with a nonionic detergent Triton X-100 combined with sonication followed by gel filtration on Sephadex G-200 and Sepharose 6B. By the data of gel filtration molecular weight of the preparation was 330,000. Two protein fractions with molecular weights of 84,000 and 90,000 were found using disc electrophoresis in 7.5% polyacrylamide gel containing 2-mercaptoethanol and sodium dodecylsulfate. The purified enzyme contained 8.6 mol of SH-groups per 100,000 daltons of protein, 4.1 micrograms of inogranic phosphate per 1 mg of protein; Km value was 37.6 microM for benzylamine as a substrate. In experiments with elective inhibitors of monoamine oxidases the enzyme from bovine heart muscle exhibited high sensitivity to the inhibitory effect of deprenyl, which characterises the monoamine oxidases of the B type.

Animals↗

PubMed1977

Significance of multiple forms of brain monoamine oxidase in situ as probed by electron spin resonance.

Spin-labeled hydroxyamphetamine, a competitive reversible inhibitor of brain monoamine oxidase, has been shown to be useful as an electron spin resonance (ESR) probe of the microenvironment of the active sites of the possible monoamine oxidase multiple forms. The ESR spectrum of spin-labeled hydroxyamphetamine was strongly quenched upon binding to the enzyme. The conformation of the active site of rat brain monoamine oxidase existing in various physical states, i.e. monoamine oxidase in situ (intact brain mitochondria), crude solubilized monoamine oxidase (MAOS) and isolated monoamine oxidase fractions (MAOa and MAOb) were critically and systematically examined. Nonlinear least squares regression analyses have been used to fit the binding data (obtained at room temperature with varying spin-labeled hydroxyamphetamine concentrations) to three groups of independent noninteracting ligand-binding models. A Gibbs-Helmholtz relationship was applied to the interpretation of the measured apparent association constant K as a function of temperature ranging from 4-50 degrees with increments of 2 degreesmfrom the extracted intensive parameters, k (intrinsic association constant) and deltaF (intrinsic free energy), as well as the apparent heat, deltaH, it was clear that the microenvironment of the binding sites existing in the more purified enzyme fractions MAOa and MAOb were similar to those found in the crude solubilized enzyme. More importantly, they correlated well with the conformation of the sites characterized in situ. The data suggested that the microenvironment of this multienzyme system was unperturbed in spite of the treatment due to the isolation process. In terms of the composition of binding sites, MAOa appeared to be heterogeneous while MAOb appeared to be more homogeneous. Since the isolated fractions MAOa and MAOb possessed marked different substrate specificities, these observations directly implied that monoamine oxidase multiple forms do exist in situ. The extracted extensive parameters, n (specific binding activity, nanomoles/mg of protein), as well as the measured characteristic transition temperatures, indicated that the relative abundance of the sites which directly affected substrate specificities was indeed altered. The consistency of the characteristic transition temperatures of 21 degrees and 38 degrees for the case of intact membrane preparations was particularly significant. A tenable hypothesis is that the manipulation in the composition of the monoamine oxidase binding forms through intimate lipid-protein interactions, which has been amply demonstrated in many biomembrane systems to be functionally important might be the underlying regulatory mechanism in vivo.

Animals↗

PubMed1973

Monoamine oxidase in rat arteries: evidence for different forms and selective localization.

1. Two forms of monoamine oxidase activity were differentiated in rat mesenteric and femoral artery by means of substrate and inhibitor specificities: one form deaminated tyramine, 5-hydroxytryptamine and noradrenaline and was highly sensitive to pargyline and clorgyline but resistant towards carbonyl reagents. This form resembled type A monoamine oxidase previously described. The other deaminated tyramine but not 5-hydroxytryptamine or noradrenaline and was inhibited by carbonyl reagents but not by clorgyline or pargyline.2. About one third of the total monoamine oxidase in homogenates of rat mesenteric artery was recovered in a 10(5)g supernatant. Both forms were partially soluble, but relatively less of the type A activity was recovered in the soluble fraction.3. Chemical sympathectomy with 6-hydroxydopamine resulted in a loss of 59% of monoamine oxidase activity in the mesenteric artery. There was a selective loss of type A activity, as revealed by the 70% decrease in 5-hydroxytryptamine deaminating ability and by the marked decrease in clorgyline sensitivity. The second monoamine oxidase species was resistant to 6-hydroxydopamine. The soluble activity was not affected by chemical sympathectomy. Most of the transmitter-specific monoamine oxidase of the arterial wall was localized within the adrenergic nerve endings. Our observations are consistent with the hypothesis that extraneuronal monoamine oxidase plays only a minor role in metabolizing noradrenaline in sympathetically innervated tissues.4. Plasma amine oxidase might originate from the arterial wall since it has similar characteristics to that found in the mesenteric artery.

Alkynes↗

PubMed1996

Novel appearance of placental nuclear monoamine oxidase: biochemical and histochemical evidence for hyperserotonomic state in preeclampsia-eclampsia.

OBJECTIVE: The aim of this study was to explore the relevance of placental monoamine oxidase at the subcellular level in the etiology of the hyperserotonomic state in preeclampsia-eclampsia. STUDY DESIGN: The study was conducted on placentas from 20 normal pregnant women and 25 women with varied severity of preeclampsia-eclampsia. Placental serotonin and subcellular monoamine oxidase activity were determined. Histochemical localization of monoamine oxidase was done in placental sections and cell isolates. RESULTS: Placental serotonin increases with severity (rsystolic 0.84, rdiastolic 0.83) and monoamine oxidase decreases (rsystolic 0.86, rdiastolic 0.79). Placental monoamine oxidase showed marked changes in preeclampsia-eclampsia. Histochemical localization of monoamine oxidase showed diffused low activity evenly throughout the cytoplasm and nucleus of the syncytiotrophoblastic cells in preeclampsia-eclampsia; in contrast, normal placenta showed high activity in the cytoplasm without any activity in the nucleus of syncytiotrophoblastic cells. Detection of monoamine oxidase activity in nuclei of the placenta in preeclampsia-eclampsia is a novel finding. Monoamine oxidase activity at the subcellular level further strengthens this observation. A severity-dependent decrease was present in the nuclei of placentas with preeclampsia-eclampsia. The use of specific substrates and inhibitors revealed the presence of monoamine oxidase in mitochondria and nucleus. CONCLUSION: The study delineates an impaired catabolism of placental serotonin in preeclampsia-eclampsia. The novel appearance of monoamine oxidase in nuclei in proximity to its normal site and low activity resulting in a hyperserotonomic state may lead to preeclampsia-eclampsia.

Adult↗

PubMed1996

Monoamine oxidase molecular activity in platelets of parkinsonian and demented patients.

Platelet monoamine oxidase activity levels have been evaluated in several neuropsychiatric disorders, to identify biochemical markers for pathological brain functioning. In the present work, we assayed both total and molecular monoamine oxidase activity in platelets of parkinsonian and demented patients: both showed significantly higher enzyme activity values than healthy controls. Thus, high platelet monoamine oxidase activity levels seem to be related to an increased intrinsic activity of single enzyme molecules. A significant positive correlation was found between platelet monoamine oxidase activity and severity of illness in both disorders: monoamine oxidase activity, therefore, may be considered as a state-dependent marker for neuro-degeneration. Such findings are discussed with reference to central nervous system biochemical abnormalities in parkinsonian and demented subjects; it might be that in both Parkinson's Disease and in dementia of Alzheimer type some central biochemical changes are reflected in certain peripheral tissues (thrombocytes, for instance), or that a systemic derangement accompanies the cerebral involvement.

Aged↗

PubMed1989

Isoelectric focusing of monoamine oxidase subtypes as identified by MAO inhibitors.

Monoamine oxidase from various human tissues from several individuals was labeled with [3H]pargyline and solubilized by means of Triton X-100 or Triton X-100 and urea. The specificity of the labeling was assessed using various selective, reversible and irreversible inhibitors as pharmacologic tools in competition experiments. The labeled material was submitted to isoelectric focusing on polyacrylamide gels according to one- and two-dimensional electrophoretic procedures and with fluorographic detection. While the differences in electrophoretic mobility of the two subtypes, MAO-A and MAO-B, could be replicated the subtypes showed identical although heterogeneous charges in isoelectric focusing. This contrasts with recent findings of clear differences in the primary structure of monoamine oxidase subtypes and thus needs further clarification.

Adult↗

PubMed2000

Monoamine oxidase inhibitors reduce conditioned fear stress-induced freezing behavior in rats.

The present study examined the acute anxiolytic effects of monoamine oxidase inhibitors on freezing behavior, a putative index of anxiety induced by conditioned fear stress. The selective serotonin 1A receptor agonist tandospirone (0.1-10 mg/kg) inhibited freezing dose dependently. The irreversible, non-selective monoamine oxidase inhibitors tranylcypromine (3 and 15 mg/kg) and phenelzine (30 and 80 mg/kg) reduced freezing significantly. Clorgyline (10 mg/kg, irreversible selective monoamine oxidase A inhibitor), N-(2-aminoethyl)-5-(m-fluorophenyl)-4-thiazole carboxamide (Ro 41-1049) (30 mg/kg, reversible selective monoamine oxidase A inhibitor), selegiline (3 mg/kg, irreversible selective monoamine oxidase B inhibitor) and lazabemide (10 mg/kg, reversible selective monoamine oxidase B inhibitor) had no effect on freezing behavior. However, combined administration of clorgyline (10 mg/kg) and selegiline (3 mg/kg) reduced freezing significantly, as well as combined administration of clorgyline (10 mg/kg) and lazabemide (10 mg/kg), Ro 41-1049 (30 mg/kg) and selegiline (3 mg/kg), or Ro 41-1049 (30 mg/kg) and lazabemide (10 mg/kg). These effects of monoamine oxidase inhibitors on freezing were not due to non-specific motor effects. These results suggest that acute inhibition of both monoamine oxidase A and B reduced anxiety or fear, while inhibition of monoamine oxidase A or B alone failed to reduce anxiety or fear.

Animals↗

PubMed1994

L-3,4-dihydroxyphenylalanine-induced dopamine release in the striatum of intact and 6-hydroxydopamine-treated rats: differential effects of monoamine oxidase A and B inhibitors.

Administration of L-DOPA (50 mg/kg) elicits a significant increase in extracellular dopamine in striata of rats treated with the catecholaminergic neurotoxin 6-hydroxydopamine but not in striata of intact rats. To assess the role of dopaminergic nerve terminals in determining the effects of exogenous L-DOPA on extracellular dopamine levels in striatum, we examined the relative contributions of monoamine oxidase A and monoamine oxidase B to the catabolism of dopamine synthesized from exogenous L-DOPA. Extracellular concentrations of dopamine and its catabolite, 3,4-dihydroxyphenylacetic acid, were monitored with in vivo dialysis in striata of intact rats and of rats with unilateral 6-hydroxydopamine lesions of striatal dopamine. Clorgyline (2 mg/kg), an inhibitor of monoamine oxidase A, significantly increased dopamine and decreased 3,4-dihydroxyphenylacetic acid in intact but not in dopamine-depleted striata. Inhibition of monoamine oxidase B with either L-deprenyl (1 mg/kg) or Ro 19-6327 (1 mg/kg) did not significantly affect dopamine or 3,4-dihydroxyphenylacetic acid in striata of intact or dopamine-depleted rats. In intact rats, administration of clorgyline in conjunction with L-DOPA produced a > 20-fold increase in dopamine and prevented the L-DOPA-induced increase in 3,4-dihydroxyphenylacetic acid. Although both L-deprenyl and Ro 19-6327 administered in combination with L-DOPA elicited a small but significant increase in dopamine, levels of 3,4-dihydroxyphenylacetic acid were not affected. In rats pretreated with 6-hydroxydopamine, clorgyline had no significant effect on the increases in dopamine and 3,4-dihydroxyphenylacetic acid elicited by L-DOPA. Furthermore, neither L-deprenyl nor Ro 19-6327 affected L-DOPA-induced increases in dopamine and 3,4-dihydroxyphenylacetic acid in dopamine-depleted striata. The present findings indicate that deamination by monoamine oxidase A is the primary mechanism for catabolism of striatal dopamine, both under basal conditions and after administration of exogenous L-DOPA. Loss of dopaminergic terminals eliminates this action of monoamine oxidase A but does not enhance deamination by monoamine oxidase B. These data support a model in which exogenous L-DOPA elicits enhanced extracellular accumulation of dopamine in the dopamine-depleted striatum because some transmitter synthesis occurs at nondopaminergic sites and the dopamine terminals that normally take up and catabolize this pool of transmitter are absent.

3,4-Dihydroxyphenylacetic Acid↗

PubMed1987

2-Chloro-2-phenylethylamine as a mechanistic probe and active site-directed inhibitor of monoamine oxidase from bovine liver mitochondria.

The reaction of 2-chloro-2-phenylethylamine with monoamine oxidase B was investigated to study the mechanism of this enzyme and its inactivation by this compound. 2-Chloro-2-phenylethylamine is a substrate with a Km of 30 microM and a turnover number of 80 min-1 at pH 6.5 at 30 degrees C. Incubation of 2-chloro-2-phenylethylamine with the enzyme led to the normal oxidation product, 2-chloro-2-phenylacetaldehyde, but only traces (0.25 mol%) of 2-phenylacetaldehyde, the product anticipated if the oxidation of substrate involved a stabilized carbanion at C-1 and elimination of chloride ion. These data suggest that a carbanion is not a likely intermediate in the oxidation of amines by monoamine oxidase. During the mechanistic studies we noted time-dependent inactivation of monoamine oxidase B by 2-chloro-2-phenylethylamine under both aerobic and anaerobic conditions. Inactivation was not reversible. Aerobically 2-chloro-2-phenylethylamine is oxidized to 2-chloro-2-phenylacetaldehyde which covalently modifies the enzyme (tau 1/2 = 40 min). Benzyl alcohol, a substrate analog, gives substantial protection against inactivation under aerobic conditions (tau 1/2 = 320 min), suggesting that an active site residue is modified. Anaerobic reaction of 2-chloro-2-phenylethylamine with monoamine oxidase B probably proceeds by direct alkylation of an enzyme residue (tau 1/2 = 140 min). Reduction with [3H]NaBH4 of the inactivated enzyme gave from 0 to 0.7 and from 4.5 to 5.6 mol of hydride incorporation for enzyme inactivated anaerobically and aerobically, respectively. The latter results are in agreement with inactivation by unmodified inhibitor and inactivation by oxidized inhibitor for the anaerobic and aerobic reactions, respectively. It is suggested that 2-chloro-2-phenylethylamine or its oxidation product 2-chloro-2-phenylacetaldehyde may serve as an active site affinity reagent for monoamine oxidase.

Aerobiosis↗