Rat Monoamine Oxidase Research Articles

Purification of Recombinant Eukaryotic MAO A and MAO B Utilizing the Pichia pastoris Expression System.

Procedures are described for the heterologous expression and purification of the mitochondrial-bound enzymes human and rat monoamine oxidases A and B and zebrafish MAO in the yeast Pichia pastoris. Enzyme expression is under control of a methanol oxidase promoter and similar procedures have been developed for the preparation of membrane particles and detergent solubilization of the functional enzymes. Similarities and differences are described in the procedures for purification of the respective enzymes using standard column chromatographic techniques to provide enzyme yields in the range of 100-300mg from 1L of cell culture.

Synthesis and monoamine oxidase B inhibitory activities of isoquiritigenin derivatives

Isoquiritigenin,one of the active constituents in the Chinese herb liquorice,is found to have moderate inhibitory activity against rat monoamine oxidase B(MAO-B,IC5047. 2 μmol·L-1). However,the structure-activity relationship(SAR) remains unclear until now. In an attempt to reveal the SAR of inhibition by isoquiritigenin,and to identify more potent and selective inhibitors of MAOB,a series of 13 derivatives based on the scaffold of isoquiritigenin were prepared,and their purities and structures were confirmed by UPLC,1 H-NMR,13 C-NMR and HRMS. These compounds were then evaluated for their ability to inhibit the enzymatic activity of human MAO-B. The SAR of inhibition was summarized and a potent compound C8 with high inhibitory activity(IC501. 4 μmol·L-1) and selectivity(>57 folds over MAO-A) was identified. Enzyme kinetics studies suggested that C8 acted as a competitive inhibitor. In addition,C8 showed little cytotoxicity to glial cells in vitro,which could be a promising lead compound for further study.

Characterization of 1-Aminobenzotriazole and Ketoconazole as Novel Inhibitors of Monoamine Oxidase (MAO): An In Vitro Investigation.

1-Aminobenzotriazole, a known time-dependent inhibitor of cytochrome P450 (CYP) enzymes, and ketoconazole, a strong inhibitor of the human CYP3A4 isozyme, are used as standard probe inhibitors to characterize the CYP and/or non-CYP-mediated metabolism of xenobiotics. In the present investigation, 1-Aminobenzotriazole and ketoconazole are characterized as potent monoamine oxidase (MAO) inhibitors in vitro using mouse, rat and human liver microsomes and S9 fractions. Inhibition potential of 1-aminobenzotriazole and ketoconazole was studied in mice, rat and human liver microsomes, S9 fractions, MAO-A and MAO-B expressed enzymes by monitoring the formation of 4-hydroxyquinoline (4-HQ) from kynuramine, a specific substrate of MAO by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Mechanism of MAO inhibition was studied by incubating varying concentration of kynuramine with mouse, rat and human S9 fractions at varying concentration of 1-aminobenzatriazole and ketoconazole and monitoring the formation of 4-HQ. 1-aminobenzotriazole and ketoconazole inhibited both MAO isozymes (MAO-A and MAO-B) with more specificity towards MAO-B. Kynuramine substrate kinetics in mouse, rat and human S9 fractions with varying 1-aminobenzotriazole and ketoconazole concentrations showed decreased maximum rate (V max) for 4-HQ formation without affecting theMichaelis-Menten constant (K m). A non-competitive inhibition model was constructed and inhibition constants (K i) for 1-aminobenzotriazole (7.87±0.61, 8.61±0.92, 65.2±1.61µM for mice, rat and humans, respectively) and ketoconazole (0.12±0.01, 2.04±0.08, 5.52±0.47µM for mice, rat and humans, respectively) were determined. 1-Aminobenzotriazole and ketoconazole are characterized as non-competitive inhibitors of mice, rat and human MAO in vitro and the extent of their MAO inhibition potential is species specific. 1-Aminobenzotriazole or ketoconazole can be used as a probe inhibitor in vitro for screening the involvement of MAO-dependent metabolism of new chemical entities (NCE) in early drug discovery.

Ciproxifan, a histamine H3 receptor antagonist, reversibly inhibits monoamine oxidase A and B

Ciproxifan is a well-investigated histamine H3 receptor (H3R) inverse agonist/antagonist, showing an exclusively high species-specific affinity at rodent compared to human H3R. It is well studied as reference compound for H3R in rodent models for neurological diseases connected with neurotransmitter dysregulation, e.g. attention deficit hyperactivity disorder or Alzheimer’s disease. In a screening for potential monoamine oxidase A and B inhibition ciproxifan showed efficacy on both enzyme isoforms. Further characterization of ciproxifan revealed IC50 values in a micromolar concentration range for human and rat monoamine oxidases with slight preference for monoamine oxidase B in both species. The inhibition by ciproxifan was reversible for both human isoforms. Regarding inhibitory potency of ciproxifan on rat brain MAO, these findings should be considered, when using high doses in rat models for neurological diseases. As the H3R and monoamine oxidases are all capable of affecting neurotransmitter modulation in brain, we consider dual targeting ligands as interesting approach for treatment of neurological disorders. Since ciproxifan shows only moderate activity at human targets, further investigations in animals are not of primary interest. On the other hand, it may serve as starting point for the development of dual targeting ligands.

Alkaloid extracts from shea butter and breadfruit as potential inhibitors of monoamine oxidase, cholinesterases, and lipid peroxidation in rats’ brain homogenates: a comparative study

Shea butter (Vitellaria paradoxa) and breadfruit (Treculia africana) are wild edible plants commonly consumed across the world. Unfortunately, these plants are currently going into extinction despite their potential biological properties. The actions of alkaloid extract from shea butter nut and breadfruit on some central nervous system biomarkers (monoamine oxidase (MAO), acetylcholinesterase (AChE), butyrylcholinesterase (BChE)) linked to some neurodegenerative diseases were investigated in this study. The radical scavenging and metal chelating abilities of the extract were also studied. Alkaloid extracts were obtained from shea butter and breadfruit via solvent extraction. The total alkaloid content of the extracts and their interactions on MAO, AChE, and BChE were determined. The ability of the extract to inhibit Fe2+-induced malondialdehyde (MDA) production in rats’ brain homogenates including their radical scavenging and metal chelating abilities were also determined. Breadfruit (17.00 mg CE/g) had higher alkaloid content than shea butter (15.34 mg CE/g). Furthermore, a significant decrease in MAO, AChE, and BChE activities was exhibited by breadfruit compared to shea butter. However, shea butter had stronger antioxidant activity. The observed modulatory effects of the extracts on MAO, AChE, and BChE activities and their antioxidant properties give credence to the fact that they could be a novel multifunctional target for the treatment and management of some neurodegenerative diseases.

In silico design of novel 2H-chromen-2-one derivatives as potent and selective MAO-B inhibitors.

Inhibition data on rat monoamine oxidase B isoform of a large number of 7-metahalobenzyloxy-2H-chromen-2-one derivatives (67 compounds) carrying at position 4 a variety of substituents differing in steric, electrostatic, lipophilic and H-bonding properties, were modeled by Gaussian field-based 3D-QSAR and docking simulations carried out on rat MAO-B homology model. The computational study combining two different approaches provided easily interpretable binding modes, highlighting the dominant role of the steric effects at position 4, and guided the design of new, potent and selective MAO-B inhibitors. The 4-hydroxyethyl-, 4-chloroethyl-, 4-carboxamidoethyl-coumarin derivatives 70, 71, and 76, respectively, were endowed with high MAO-B inhibitory potency (pIC50=8.13, 7.89 and 7.82, respectively) and good selectivity over MAO-A (pIC50=5.33, 3% inhibition at 10μM, and pIC50=5.37, respectively). New compounds with moderate to low MAO-B inhibitory activity were also designed and prepared to challenge the predictive power of our docking-based 3D-QSAR model. The good match between predicted and experimental pIC50 values for all the newly designed compounds confirmed the robustness of our model (r(2)=0.856, RMSE=0.421) and its transparent rationale in unveiling the main molecular determinants for high potency towards MAO-B.

Acción Anticonvulsivante de los Inhibidores de la Monoamino Oxidasa

Para la evaluación de fármacos anticonvulsivantes, se han usado una serie de métodos experimentales con los más variados resultados. Una de las técnicas que ha demostrado buena correlación con la eficacia clínica anticonvulsiva está basada en la habilidad de ciertos fármacos para alterar el carácter de la crisis producida por el electroshock supramóximo (12). El hecho de que los anticonvu1sivantes clásicos: barbitúricos e hidantoínas, ejerzan sus efectos protectores anticonvulsivantes, acompañados de sedación del sistema nervioso central, ha mantenido la inquietud de los investigadores por encontrar el anticonvulsivante ideal. Estudios realizados por Prockop et al. en 1959, han mostrado que los inhibidores de la monoaminoxidasa (IMAO) evitan en las ratas la presentación de la fase de extensión tónica del tren posterior, producida por el choque eléctrico supramáximo. A diferencia de los anticonvuIsivantes clásicos los inhibidores de la monoaminoxidasa poseen cualidades psicoanalépticas (29).

2H Kinetic Isotope Effects and pH Dependence of Catalysis as Mechanistic Probes of Rat Monoamine Oxidase A: Comparisons with the Human Enzyme

Monoamine oxidase A (MAO A) is a mitochondrial outer membrane-bound flavoenzyme important in the regulation of serotonin and dopamine levels. Because the rat is extensively used as an animal model in drug studies, it is important to understand how rat MAO A behaves in comparison with the more extensively studied human enzyme. For many reversible inhibitors, rat MAO A exhibits K(i) values similar to those of human MAO A. The pH profile of k(cat) for rat MAO A shows a pK(a) of 8.2 ± 0.1 for the benzylamine ES complex and pK(a) values of 7.5 ± 0.1 and 7.6 ± 0.1 for the ES complexes with p-CF(3)-(1)H- and p-CF(3)-(2)H-benzylamine, respectively. In contrast to the human enzyme, the rat enzyme exhibits a single pK(a) value (8.3 ± 0.1) with k(cat)/K(m) for benzylamine versus pH and pK(a) values of 7.8 ± 0.1 and 8.1 ± 0.2 for the ascending limbs, respectively, of k(cat)/K(m) versus pH profiles for p-CF(3)-(1)H- and p-CF(3)-(2)H-benzylamine and 9.3 ± 0.1 and 9.1 ± 0.2 for the descending limbs, respectively. The oxidation of para-substituted benzylamine substrate analogues by rat MAO A has large deuterium kinetic isotope effects on k(cat) and on k(cat)/K(m). These effects are pH-independent and range from 7 to 14, demonstrating a rate-limiting α-C-H bond cleavage step in catalysis. Quantitative structure-activity correlations of log k(cat) with the electronic substituent parameter (σ) at pH 7.5 and 9.0 show a dominant contribution with positive ρ values (1.2-1.3) and a pH-independent negative contribution from the steric term. Quantitative structure-activity relationship analysis of the binding affinities of the para-substituted benzylamine analogues for rat MAO A shows an increased van der Waals volume (V(w)) increases the affinity of the deprotonated amine for the enzyme. These results demonstrate that rat MAO A exhibits functional properties similar but not identical with those of the human enzyme and provide additional support for C-H bond cleavage via a polar nucleophilic mechanism.

Interactions of Monoamine Oxidases with the Antiepileptic Drug Zonisamide: Specificity of Inhibition and Structure of the Human Monoamine Oxidase B Complex

The binding of zonisamide to purified, recombinant monoamine oxidases (MAOs) has been investigated. It is a competitive inhibitor of human MAO B (K(i) = 3.1 ± 0.3 μM), of rat MAO B (K(i) = 2.9 ± 0.5 μM), and of zebrafish MAO (K(i) = 30.8 ± 5.3 μM). No inhibition is observed with purified human or rat MAO A. The 1.8 Å structure of the MAO B complex demonstrates that it binds within the substrate cavity.

High-level expression and purification of rat monoamine oxidase A (MAO A) in Pichia pastoris: Comparison with human MAO A

The high-level heterologous expression in Pichia pastoris, purification and characterization of recombinant membrane-bound rat liver monoamine oxidase A (MAO A) are described. A 1-L culture of cells produces approximately 700 U of rat MAO A activity. The rat MAO A activity is found in outer mitochondrial membrane of the cell. Using a modification of the human MAO A purification procedure, approximately 200mg of recombinant rat MAO A is purified in a 43% yield and exhibits a molecular weight of approximately 60,000 kDa on SDS-PAGE. The purified enzyme contains a covalently bound FAD and forms a N(5) flavocyanine adduct on inhibition by clorgyline. Edman sequencing shows that the amino terminus of rat MAO A is blocked at an N-terminal threonyl residue. The purified rat enzyme exhibits a higher thermal stability than does purified human MAO A. Compared with human MAO A, rat MAO A oxidizes serotonin or kynuramine with twofold higher k(cat)/K(m) values, oxidizes phenethylamine with a 6.7-fold higher catalytic efficiency and benzylamine with a approximately 40-fold higher catalytic efficiency. Although approximately 90% identical in sequence to human MAO A, rat MAO A is a more efficient catalyst for amine neurotransmitter oxidation.

Characterization of detergent purified recombinant rat liver monoamine oxidase B expressed in Pichia pastoris

The high level expression and purification of rat monoamine oxidase B (rMAOB) in the methylotrophic yeast Pichia pastoris is reported. Nearly 100 mg of purified rMAOB is obtained from 130 g (wet weight) of cells (0.5 L of culture). The MALDI-TOF mass spectrum of the purified protein shows a single species with a molecular mass of 59.228 ± 0.064 kDa, which agrees with the calculated molecular weight of 59.172 kDa for the rMAOB protein sequence assuming one mole of covalent FAD per mole of the enzyme. Consistent with the MALDI-MS data, purified rMAOB shows a single band near 60 kDa in Coomassie-stained SDS–PAGE gel as well as on Western blot analyses performed using antisera raised against human MAOA and BSA-conjugated FAD. A partial amino acid sequence of the purified protein is confirmed to be that of the wild type rMAOB by in-gel trypsin digestion and MALDI-TOF-MS analyses of the liberated peptide fragments. Steady state kinetic data show that purified rMAOB exhibits a K m(amine) of 176 ± 15 μM and a k cat of 497 ± 83 min −1 for benzylamine oxidation, and a K m(O 2) of 170 ± 10 μM. Kinetic parameters obtained for purified rMAOB are compared with those reported earlier for recombinant human liver MAOB expressed in P. pastoris.

Determination of the Oligomeric States of Human and Rat Monoamine Oxidases in the Outer Mitochondrial Membrane and Octyl β-d-Glucopyranoside Micelles Using Pulsed Dipolar Electron Spin Resonance Spectroscopy

Human monoamine oxidase A (hMAOA) is considered to be unique among mammalian MAOs in having a non-conservative Glu-X-Lys mutation (X being 151 in MAOAs and 142 in MAOB's), which is suggested to be the reason for its monomeric structure. This hypothesis has been tested in this work. A pargyline based nitroxide spin labeled irreversible inhibitor (ParSL) was used as a MAO active site specific spin probe to measure intersubunit distances in detergent (octyl beta-d-glucopyranoside, OGP) purified and OMM bound forms by a pulsed dipolar ESR spectroscopic (PDS) technique. In a parallel approach, the covalent flavin cofactor present in the MAO active sites was reduced to its respective anionic flavin semiquinone and used for measuring inter-flavin distances in detergent purified samples. The measured interspin distances are within 0.1-0.3 nm of those estimated from the available dimeric crystal structures of human MAOB and rat MAOA and show that all human and rat MAOs exist as dimers in the OMM. In the OGP micelle, however, human and rat MAOAs exist only partially (<or=50%) as dimers, whereas human and rat MAOBs exist entirely as dimers. The Lys-151-Glu mutant of human MAOA and the Glu-142-Lys mutant of human MAOB exhibit similar spectral properties as the corresponding wild-type enzymes. Therefore, no role of the Glu-X residue in stabilizing dimeric structures of MAOs was found. The monomeric crystal structure reported for human MAOA is thus a result of its instability in the OGP micelles. The general applicability of the PDS technique to structural studies of membrane proteins in their native membrane environments and detergent purified forms is discussed.

Comparison of the Structural Properties of the Active Site Cavities of Human and Rat Monoamine Oxidase A and B in Their Soluble and Membrane-Bound Forms

Structural properties of the active site cavities in human and rat monoamine oxidases (MAOA and MAOB) have been studied in their detergent-purified and outer mitochondrial membrane (OMM) bound forms using a spin-labeled irreversible inhibitor (ParSL) as an active specific spin probe. ParSL has been found to be 5-10-fold more specific for human MAOB (hMAOB) with a Ki of ca. 20 muM, compared to Ki's in the range of 100-200 muM observed for other human and rat MAOs. Solvent accessibilities of the active-site-bound spin probes have been determined by studying the power saturation properties of the spin probe EPR signals in the presence and absence of a polar paramagnetic reagent NiEDDA and by measuring the extent of spin probe reductions on treatment with excess ascorbic acid. Results presented here show that the spin probe bound to the hMAOA active site is ca. 7-8-fold more accessible than in hMAOB. In contrast, the spin probes covalently attached to the two rat enzyme active sites show comparable accessibilities to each other. On comparison of human versus rat enzymes, the active-site-bound spin probes in the two rat MAOs show ca. 40% less accessibilities compared to the same in hMAOA but ca. 4-5-fold higher accessibilities than in hMAOB active site. The present data thus suggests that the structural properties of the active site cavities in rat MAOs are significantly different compared to those in the two human enzymes, which correlates with the differences reported earlier in the inhibitor specificities between human and rat MAOs.

Molecular characterization of monoamine oxidase in zebrafish ( Danio rerio)

Monoamine oxidase (MAO) is responsible for the degradation of a number of neurotransmitters and other biogenic amines. In terrestrial vertebrates, two forms of the enzyme, named MAO A and B, were found in which mammals are coded by two similar but distinct genes. In teleosts, the biochemical data obtained so far indicate that enzyme activity is due to a single form, whose sequence, obtained for trout, displays 70% identity with mammal MAO A and B. In this paper, we carried out an investigation of zebrafish MAO (Z-MAO) to shed further light on the nature of the MAO form present in aquatic vertebrates. Sequencing studies have revealed an open reading frame 522-amino-acids long with MW 58.7 kDa, displaying 84% identity with trout MAO and about 70% identity with mammal MAO A and MAO B. Analysis of the sequence and of the predicted secondary structure shows that also in Z-MAO principal domains characterizing the MAOs are present. The domain linking the FAD is very well conserved, while the transmembrane domain sequence linking the enzyme to the external mitochondrial membrane does not appear to be conserved even with respect to trout MAO. Comparison with the amino acids which, according to the human MAO B and rat MAO A models, line the substrate-binding site shows that in Z-MAO, several residues (V172, N173, F200, L327) differ from MAO B but are similar or identical to the corresponding ones present in rat MAO A, as well as in trout MAO. A three-dimensional model is reported of the substrate-binding site of Z-MAO obtained by comparative modeling. Our observations support the hypothesis that the MAO form present in aquatic vertebrates is a MAO A-like form. Experiments performed to test the effect of selective MAO A (clorgyline) and MAO B (deprenyl) inhibitors on the enzyme's activity in liver and brain confirm the presence of a single form of MAO in zebrafish.

Decreases in Glucocorticoid Sensitivity as a Factor of Stress-Producing Changes in the Activity of Monoamine Oxidase, Lipid Peroxidation, and Behavior in Rats

Four episodes of immobilization stress induced a decrease in the sensitivity of rats to glucocorticoid hormones, which was accompanied by anxiogenic behavior, increased MAO-B activity, and a parallel increase in lipid peroxidation (LPO) in brain tissues. There was a simultaneous increase in MAO-B activity in the kidneys and accumulation of LPO products in the liver and kidneys. Administration of Kenalog (2 mg/kg), a pharmacological analog of glucocorticoid hormones, prevented the poststress activation of MAO-B and LPO and decreased the extent of anxiogenic behavioral abnormalities in rats.

Structure of Rat Monoamine Oxidase A and Its Specific Recognitions for Substrates and Inhibitors

Monoamine oxidase (MAO), a mitochondrial outer membrane enzyme, catalyzes the degradation of neurotransmitters in the central nervous system and is the target for anti-depression drug design. Two subtypes of MAO, MAOA and MAOB, are similar in primary sequences but have unique substrate and inhibitor specificities. The structures of human MAOB complexed with various inhibitors were reported early. To understand the mechanisms of specific substrate and inhibitor recognitions of MAOA and MAOB, we have determined the crystal structure of rat MAOA complexed with the specific inhibitor, clorgyline, at 3.2Å resolution. The comparison of the structures between MAOA and MAOB clearly explains the specificity of clorgyline for MAOA inhibition. The fitting of serotonin into the binding pockets of MAOs demonstrates that MAOB Tyr326 would block access of the 5-hydroxy group of serotonin into the enzyme. These results will lead to further understanding of the MAOA function and to new anti-depression drug design. This study also presents that MAOA has a transmembrane helix at the C-terminal region. This is the first crystal structure of membrane protein with an isolated transmembrane helix.

Crystallization and preliminary crystallographic analysis of rat monoamine oxidase A complexed with clorgyline.

Monoamine oxidase (MAO) is an FAD-containing mitochondrial outer-membrane protein which catalyzes the degradation of several neurotransmitters in the central nervous system. The two subtypes of MAO, MAOA and MAOB, have similar primary sequences but different substrate and inhibitor specificities. The structure of human MAOB has recently been determined, but the structure of MAOA remains unknown. To clarify the mechanisms underlying their unique substrate and inhibitor recognition and thereby facilitate the development of new specific inhibitors to treat MAO-related neurological disorders, rat MAOA was crystallized in a complex with the specific inhibitor clorgyline. Diffraction data were collected to 3.2 A resolution. The crystal belongs to the space group P4(3)2(1)2, with unit-cell parameters a = b = 158.2, c = 258.4 A.

Decreased glucorticoid sensitivity as a factor of stressogenic shifts in the activity of monoamine oxidase, lipid peroxidation, and behavior in rats

Four episodes of immobilization stress cause a decrease in the sensitivity to glucocorticoid hormones, followed by anxiogenic be­havioral disorders, enhanced monoamine oxidase-В (МАО-В) activity and simultaneously increased lipid peroxidation (LPO) in the brain tissue of rats. Concurrently, there is an increase in renal МАО-В activity, as well as renal and hepatic accumulation of LPO products. Administration of kenalog (2 mg/kg), a phar­macological analogue of glucocorticoid hormones, prevents the poststress МАО-В activation and LPO and attenuates anxiogen­ic behavioral disorders in the rats.

Phe(208) and Ile(199) in human monoamine oxidase A and B do not determine substrate and inhibitor specificities as in rat.

It has been reported previously that reciprocally switching Phe(208) and Ile(199) in rat monoamine oxidase (MAO) A and B, respectively, was sufficient to switch their substrate and inhibitor preferences. In this study, the same mutants were made in the human forms of MAO. When compared with MAO A, MAO A-F208I showed a sixfold decrease in the specificity constant k(cat)/K(m) for both the MAO A- and the MAO B-preferring substrates 5-hydroxytryptamine and beta-phenylethylamine, respectively. The reciprocal point mutant MAO B-I199F had no effect on substrate affinity. To investigate if the region neighboring these two residues is responsible for conferring preferences, we have also made chimeric constructs by reciprocally switching the corresponding amino acid segments 159-214 in MAO A and 150-205 in MAO B. Chimerics MAO AB(159-214)A and MAO BA(150-205)B had small changes in K(m) and IC(50) values when compared with MAO A and B, respectively, but did not exhibit a preference switch. The results suggest that Phe(208) in MAO A and amino acid segments 159-214 and 150-205 in MAO A and B, respectively, influence the enzyme active site. However, substrate and inhibitor preferences of human MAO A and B are not determined by the respective residues Phe(108) and Ile(199) as in rat MAO nor by their neighboring regions.

In vitro biotransformation of the selective serotonin reuptake inhibitor citalopram, its enantiomers and demethylated metabolites by monoamine oxidase in rat and human brain preparations.

This study was conducted to identify enzyme systems eventually catalysing a local cerebral metabolism of citalopram, a widely used antidepressant of the selective serotonin reuptake inhibitor type. The metabolism of citalopram, of its enantiomers and demethylated metabolites was investigated in rat brain microsomes and in rat and human brain mitochondria. No cytochrome P-450 mediated transformation was observed in rat brain. By analysing H2O2 formation, monoamine oxidase A activity in rat brain mitochondria could be measured. In rat whole brain and in human frontal cortex, putamen, cerebellum and white matter of five brains monoamine oxidase activity was determined by the stereoselective measurement of the production of citalopram propionate. All substrates were metabolised by both forms of MAO, except in rat brain, where monoamine oxidase B activity could not be detected. Apparent Km and Vmax of S-citalopram biotransformation in human frontal cortex by monoamine oxidase B were found to be 266 microM and 6.0 pmol min(-1) mg(-1) protein and by monoamine oxidase A 856 microM and 6.4 pmol min(-1) mg(-1) protein, respectively. These Km values are in the same range as those for serotonin and dopamine metabolism by monoamine oxidases. Thus, the biotransformation of citalopram in the rat and human brain occurs mainly through monoamine oxidases and not, as in the liver, through cytochrome P-450.