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.     

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.     

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     

Some factors influencing the metabolism of benzylamine by type A and B monoamine oxidase in rat heart and liver

The ability of MAO-A and MAO-B to metabolize benzylamine in vitro has been investigated in mitochondrial preparations from rat liver and heart. Although under normal circumstances benzylamine appeared to be metabolized exclusively by MAO-B in the rat liver, a contribution by both MAO-A and a clorgyline-resistant enzyme component was revealed when the MAO-B activity was much reduced by pretreatment of the mitochondria with appropriate concentrations of deprenyl. These three enzyme activities also contributed to benzylamine deamination in rat heart mitochondria. However, binding studies with [3H]pargyline, which provided an estimate of the respective concentrations of MAO-A and MAO-B active centres in heart mitochondria, indicated a ratio between MAO-A and MAO-B, markedly different from that shown by plots of inhibition of benzylamine metabolism by various concentrations of clorgyline. The interpretation of these clorgyline plots is discussed in terms of the kinetic constants of both MAO-A and MAO-B, and the relative amounts of each enzyme. It is proposed that although the turnover rate constant for benzylamine metabolism by MAO-A is much smaller than that shown by MAO-B, in those tissues containing a large ratio of MAO-A:MAO-B content, the metabolism of benzylamine by MAO-A can be detected.     

Some factors influencing the metabolism of benzylamine by type A and B monoamine oxidase in rat heart and liver

The ability of MAO-A and MAO-B to metabolize benzylamine in vitro has been investigated in mitochondrial preparations from rat liver and heart. Although under normal circumstances benzylamine appeared to be metabolized exclusively by MAO-B in the rat liver, a contribution by both MAO-A and a clorgyline-resistant enzyme component was revealed when the MAO-B activity was much reduced by pretreatment of the mitochondria with appropriate concentrations of deprenyl. These three enzyme activities also contributed to benzylamine deamination in rat heart mitochondria. However, binding studies with [³ H]pargyline, which provided an estimate of the respective concentrations of MAO-A and MAO-B active centres in heart mitochondria, indicated a ratio between MAO-A and MAO-B, markedly different from that shown by plots of inhibition of benzylamine metabolism by various concentrations of clorgyline. The interpretation of these clorgyline plots is discussed in terms of the kinetic constants of both MAO-A and MAO-B, and the relative amounts of each enzyme. It is proposed that although the turnover rate constant for benzylamine metabolism by MAO-A is much smaller than that shown by MAO-B, in those tissues containing a large ratio of MAO-A:MAO-B content, the metabolism of benzylamine by MAO-A can be detected.     

Monoamine oxidase activities in lymphocytes and granulocytes taken from pig blood

The characterisation of monoamine oxidase activities in lymphocytes and granulocytes was studied using cells prepared from pig blood. The specific activities against beta-phenylethylamine, benzylamine, tyramine and 5-hydroxytryptamine as substrates in granulocytes (G) were approximately twice those found in lymphocytes (L). The absence of the semicarbazide-sensitive amine oxidase (SSAO) was confirmed by insensitivity of the latter to semicarbazide as inhibitor with benzylamine as substrate. MAO activity present in (G) and (L) was selectively inhibited by low deprenyl concentrations; this fact, in addition to the simple sigmoid inhibition curves obtained with increasing concentrations of clorgyline with tyramine as substrate, suggests that the MAO activity present both in (G) and (L) is predominantly of the MAO-B form. The absence of any contamination with plasma amine oxidase (EC 1.4.3.6) was confirmed by the fact that activity towards benzylamine (Bz) was insensitive to KCN-induced inhibition. Kinetic constants were determined for each fraction towards beta-phenylethylamine (PEA) and Bz as substrates. MAO-B was titrated with unlabelled pargyline, deprenyl and [3H]-pargyline; the corresponding Kcat values, turnover number and the active concentrations were then determined. The molecular weight of MAO-B present in both cellular fractions was calculated by SDS-electrophoresis and fluorography, after reaction with [3H]-pargyline. Some of these results are compared with those obtained with human blood leucocytes.     

Characterization of N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) binding sites in C57BL/6 mouse brain: mutual effects of monoamine oxidase inhibitors and sigma ligands on MPTP and sigma binding sites.

N-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induces Parkinson-like symptoms in humans, nonhuman primates, and mice. Several studies suggest that MPTP is metabolized by monoamine oxidase (MAO) type B to yield N-methyl-4-phenyl-pyridinium (MPP+), which is responsible for the neurotoxic effects of the drug. In the present study, the pharmacological properties of [3H]MPTP binding sites in C57BL/6 mouse brain membranes were investigated, and a possible relationship to the sigma binding sites was examined. Both equilibrium binding experiments and kinetic assays indicate that [3H]MPTP labels two distinct binding sites in C57BL/6 mouse brain. The high affinity [3H]MPTP binding sites (Kd = 13 nM) are selectively blocked by the MAO type A inhibitor clorgyline, and the residual low affinity [3H]MPTP sites (Kd = 1100 nM) display the pharmacological specificity of MAO-B binding sites. In contrast, the low affinity [3H]MPTP binding sites are blocked by the selective MAO-B inhibitor (-)-deprenyl, and the drug-specificity profile of the remaining high affinity sites is consistent with the properties of MAO-A binding sites. The affinities of several MAO inhibitors tested and of MPTP for the high affinity MPTP/MAO-A binding sites correlate well (r = 0.96) with their affinities for the sigma binding sites labeled with [(+)-[3H]-3-PPP]. The sigma receptor ligand (+)-3-PPP displays moderately high affinity for the MPTP/MAO-A binding sites but negligible affinity for MPTP/MAO-B sites. Moreover, (+)-3-PPP alters the dissociation kinetics of MPTP from the high affinity MPTP/MAO-A sites. The finding that [3H]MPTP labels MAO-B sites supports the hypothesis that the drug is a substrate for these enzyme binding sites. However, the finding that the high affinity sites, labeled by [3H] MPTP, are particularly sensitive to MAO-A inhibitors, which also display high affinity for the sigma binding sites, may suggest a possible relationship between MAO-A and sigma binding sites. In turn, the kinetic experiments imply that sigma ligands [i.e., (+)-3-PPP] may allosterically modulate the binding to MAO-A binding sites.     

The effects of selective and non-selective monoamine oxidase (MAO) inhibitors on conflict behavior in the rat

Conflict behavior in rats was examined over the course of several weeks of chronic treatment with selective and non-selective monoamine oxidase inhibitors (MAOIs). In daily 10min sessions, rats were trained to drink from a tube which was occasionally electrified (0.5mA). Electrification was signalled by the presence of a tone. Within 3-4 weeks, control (i.e. non-drug) conflict behavior had stabilized (30-40 shocks and 8-12ml water/session) and drug testing began. Chronic administration (two injections/day for 8 weeks) with a non-selective (i.e. MAO-A and MAO-B inhibiting) dose of pargyline (15mg/kg) resulted in a time-dependent increase in punished responding. In contrast, chronic administration of the MAO-A selective inhibitor (clorgyline; 1.0mg/kg, 2mg/kg), the MAO-B selective inhibitor deprenyl (5mg/kg) or MAO-B inhibiting doses of pargyline (1.0mg/kg, 5mg/kg) were without effect. Finally, chronic treatment with the combination of a low dose of clorgyline (1.0mg/kg) and a low dose of pargyline (1.0mg/kg) did result in a time-dependent increase in punished responding. These results suggest that inhibition of both MAO-A and MAO-B is required for the eventuation of the anxiolytic effect resulting from chronic MAOI treatment.     

Synthesis, molecular modeling studies, and selective inhibitory activity against monoamine oxidase of 1-thiocarbamoyl-3,5-diaryl-4,5-dihydro-(1H)- pyrazole derivatives

A novel series of 1-thiocarbamoyl-3,5-diaryl-4,5-dihydro-(1H)-pyrazole derivatives have been synthesized and investigated for the ability to inhibit selectively the activity of the A and B isoforms of monoamine oxidase (MAO). All the synthesized compounds show high activity against both the MAO-A and the MAO-B isoforms with Ki values between 27 and 4 nM and between 50 and 1.5 nM, respectively, except for a few derivatives whose inhibitory activity against MAO-B was in the micromolar range. Knowing that stereochemistry may be an important modulator of biological activity, we performed the semipreparative chromatographic enantioseparation of the most potent, selective, and chiral compounds. The separated enantiomers were then submitted to in vitro biological evaluation. The selectivity of the (-)-(S)-1 enantiomer against MAO-B increases twice and a half, while the selectivity of the (-)-(S)-4 enantiomer against MAO-A triples. Both the MAO-A and MAO-B isoforms respectively of the 1O5W and 1GOS models deposited in the Protein Data Bank were considered in the computational study. The docking study was carried out using several computational approaches with the aim of proposing possible binding modes of the MAO enantioselective compounds 1 and 4.     

Inhibition of MAO-A activity enhances behavioural activity of rats assessed using water maze and open arena tasks.

To determine if the inhibition of MAO-A and/or MAO-B activities can influence cognitive processes in adult rats, we analysed whether chronic treatment with clorgyline, 1-deprenyl and pargyline could modify the performance of adult rats in a modified version of the water maze task. The effects of these treatments on locomotor activity and enzyme activities were also assessed. Rats were treated for 24 days with clorgyline (0.2 mg/kg), 1-deprenyl (0.25 mg/kg) and pargyline (I or 10 mg/kg). The treatments were started two weeks before the water maze experiment and continued until the end of testing. The rats were trained to find a submerged platform (6 days: I trial/day; 7 th day: probe trial). Over the next three days, locomotor activity was assessed in an open arena. Treatments with clorgyline (MAO-A inhibitor), 1-deprenyl (MAO-B inhibitor) and pargyline (non-selective MAO inhibitor) did not improve the finding of the hidden platform, when compared to treatment with saline, but significantly increased the swimming speed of the rats. The different treatments, when compared to saline, failed to modify the distance covered and the number of groomings performed in the open arena. However, clorgyline and pargyline, 10 mg/kg, increased the number of faecal boli and clorgyline enhanced the number of rearings made when compared to saline, 1-deprenyl and pargyline, 10 mg/kg. These results indicate that near total inhibition of MAO-A by clorgyline and pargyline as assessed by MAO activity measurement induces an increase in locomotor activity but that inhibition of MAO-A or MAO-B, either alone or combined, does not facilitate spatial learning in adult rats.     

Effect of a new antiparkinsonian drug himantane on monoamine oxidase activity

N-2(adamantyl)hexamethyleneimine hydrochloride (hemantane) is a new potential antiparkinsonian drug targeted at several neurochemical systems. The drug exhibits the properties of a low-affinity noncompetitive blocker of the ion channels of glutamate NMDA receptors. Hemantane increases the content of dopamine in the striatum, while decreasing the level of dopamine metabolite dioxyphenylacetic acid (DOPAC). Investigation of the drug interaction with monoamine oxidases (MAOs) of the A and B types in vitro showed that hemantane acts as a weak competitive inhibitor of MAO-B (Ki = 470 +/- 70 microM) and partly protected MAO-B from irreversible inhibition by selegiline (deprenyl), while virtually not influencing the activity of MAO-A. Administered to C57BL6 mice (20-100 mg/kg), hemantane did not influence the activity of MAO-B measured in isolated cerebral mitochondria. At the same time, hemantane administered in combination with deprenyl significantly reduced activity of the latter drug and caused pronounced irreversible inhibition of mitochondrial MAO-B (comparable with the effect of deprenyl introduced alone). Therefore, inhibition of MAO-B may contribute to the spectrum of neurochemical activity of hemantane.     

Stereospecific deuterium substitution at the alpha-carbon position of dopamine and its effect on oxidative deamination catalyzed by MAO-A and MAO-B from different tissues

Stereospecific replacement of deuterium in the alpha-carbon side chain position of dopamine (DA) was achieved by decarboxylation of L-3,4-dihydroxyphenylalanine (L-dopa) using hog kidney aromatic aminoacid decarboxylase. The S[alpha-2H1]DA enantiomer was obtained by decarboxylation of L-[alpha-2H1]dopa in H2O, while the R[alpha-2H1]DA enantiomer was obtained by decarboxylation of unsubstituted L-dopa in 2H2O. An inverse solvent isotope effect of L-dopa decarboxylation was observed in 2H2O. The deaminated aldehyde products of the four DA analogues, i.e. undeuterated DA, [alpha, alpha-2H2] DA, R[alpha-2H1]DA and S[alpha-2H1]DA, have been analyzed by the gas chromatography-mass spectrometry (GC-MS) method. It is evident that monoamine oxidase (MAO) catalyzes the stereochemical removal of only R-deuterium and that S-deuterium was maintained at the alpha-carbon atom of 3,4-dihydroxyphenylacetaldehyde. The steady-state kinetics of the oxidative deamination of undeuterated, [alpha, alpha-2H2], R[alpha-2H1], and S[alpha-2H1] dopamine were assessed by determination of the aldehyde products directly by high performance liquid chromatography (HPLC) using electrochemical detection (ECD). MAO-A from rat liver mitochondria (deprenyl-treated) and from human placenta, as well as MAO-B from rat liver (clorgyline-treated) and from human platelet were used in this study. The apparent isotope effects, i.e. (V/K)H/(V/K)D ratios of [alpha, alpha-2H2]DA and R[alpha-2H1]DA, were quite similar (2.34 and 3.13) with respect to both MAO-A and MAO-B. S[alpha-2H1]DA exhibited a slight secondary isotope effect. Formula: see text.     

The neuropharmacological profile of N-methyl-N-propargyl-2-aminotetralin: a potent monoamine oxidase inhibitor

Summary N-methyl-N-propargyl-2-aminotetralin (N-0425), a semi rigid analogue of deprenyl was found to be a potent inhibitor of monoamine oxidase type-A and-B. The MAO inhibitory potency was determined in in-vitro, ex-vivo and in-vivo experiments for racemic N-0425 and for both enantiomers, and compared with deprenyl. Racemic N-0425 and (–)N-0425 were found to inactivate both MAO-A and-B to about the same extent in rat brain homogenates, whereas (+)N-0425 was 10 times more potent in inhibiting MAO-B than MAO-A under in-vitro conditions. The latter compound was almost 3 times less active than (–)deprenyl with respect to inactivation of MAO-B. In ex-vivo experiments it was shown that (±)- and (+)N-0425 inhibited rat striatal MAO-B activity almost completely 2 h after a dose of 0.01 mmol/kg ip, whereas both compounds produced a much less rapid inactivation of type-A MAO, which was about 65% after 23 h. No potentiation of the tyramine induced increase in systolic blood pressure was found in normotensive rats following doses up to 0.01 mmol/kg ip of racemic N-0425, but a potentiation was observed after a higher dose of 0.04 mmol/kg. Levels of dopamine and noradrenaline were both increased in rat frontal cortex after the administration of N-0425, which can be interpreted as a reflection of MAO inactivation. Since we were able to select a dose of (±)N-0425 which potently inhibits MAO, without producing a concomittant potentiation of the tyramine effect on blood pressure in normotensive rats, it is reasonable to suggest that this compound, like deprenyl, could be useful in the treatment of Parkinson's disease.Abbreviations used MAO monoamine oxidase - l-DOPA 3,4-dihydroxyphenylalamine - DA dopamine - DOPAC 3,4-dihydroxyphenylacetic acid - HVA homovanillic acid - 3-MT 3-methoxytyramine - NA noradrenaline - NM normetanephrine - 5-HT 5-hydroxytryptamine - 5-HIAA 5-hydroxyindoleacetic acid - PEA -phenylethylamine - N-0425 N-methyl-N-propargyl-2-aminotetralin Some of these results have been presented in a preliminary form at the 5th International Catecholamine Symposium, Göteborg, Sweden, June 12–16, 1983. This work was partly supported by a grant from Nelson Research, Irvine, California, USA     

Inhibition of rat brain monoamine oxidase activity by cerebral anti-ischemic agent, ifenprodil

Ifenprodil, which is clinically used as a cerebral vasodilator, inhibited rat brain type A (MAO-A) and type B (MAO-B) monoamine oxidase activity. It did not, however, affect rat lung semicarbazide-sensitive amine oxidase. The degree of inhibition of either form of MAO was not changed by 30 min preincubation of the enzyme preparations at 37 degrees C with ifenprodil. Modes of inhibition of MAO-A and MAO-B by ifenprodil were competitive towards oxidation of their respective substrates, 5-hydroxytryptamine and benzylamine, with Ki values of 75 microM for inhibition of MAO-A and 110 microM for inhibition of MAO-B.     

Irreversible Inhibition of Rat Liver Mitochondrial MAO A and MAO B by Enantiomers of Deprenyl and α-Methylpargyline

Using rat liver mitochondrial monoamine oxidase (MAO) A and MAO B, the possible influence of stereochemical factors upon the irreversible inhibition by propargylamine derivatives has been studied using the enantiomers of deprenyl and of α-methylpargyline. Whether studying the inhibition of MAO A or MAO B, little difference was found among enantiomeric pairs in the first-order rate constant (k₂) for formation of the enzyme inhibitor adduct. Similarly, and with the exception of (S)-d -(+)-deprenyl (k₂ = 0 or an extremely low value at MAO A), the computed value of k₂ for the individual enantiomers showed little variation between MAO A and MAO B. These results suggest that inhibitor selectivity towards a particular form of the enzyme is determined predominantly at the competitive phase of the inhibition.     

Stereochemical effects of 11-OH-delta 8-tetrahydrocannabinol-dimethylheptyl to inhibit adenylate cyclase and bind to the cannabinoid receptor

The recent preparation of the enantiomers of 11-OH-delta 8-tetrahydrocannabinol-dimethylheptyl (THC-DMH), recrystallized to absolute enantiomeric purity, has made it possible to examine the requirement for stereospecificity for the interaction of this component with the cannabinoid receptor, defined by the binding of [3H]CP-55,940 and the adenylate cyclase enzyme. The enantiomer (-)11-OH-delta 8-THC-DMH exhibited a fully efficacious and potent (IC50 = 1.8 nM) inhibition of the accumulation of cAMP in intact N18TG2 cells. The (-)enantiomer was as efficacious and potent (Kinh = 7.2 nM) as desacetyllevonantradol in inhibiting adenylate cyclase activity in membrane preparations. The (-)enantiomer was able to compete fully for the specific binding of [3H]CP-55,940 to membranes from the brain of the rat in homologous displacement studies (Ki = 234 pM). The potency ratios exhibited by the (-) to (+)enantiomers of 11-OH-delta 8-THC-DMH exceeded 1000 for each of these activities.     

Enantiomer/enantiomer interaction of (S)- and (R)-propafenone for cytochrome P450IID6-catalyzed 5-hydroxylation: in vitro evaluation of the mechanism

Many drugs are used as racemates, and the enantiomers may differ in terms of pharmacological properties and disposition. Stereoselective disposition of the enantiomers can arise from metabolism of the enantiomers via different routes catalyzed by different enzymes. In contrast, the enantiomers may be metabolized by the same enzyme at different rates. In the latter case, the enantiomers can compete for this metabolic step, giving rise to the possibility of an enantiomer/enantiomer interaction. We have chosen the antiarrhythmic propafenone, for which in vivo data indicated an interaction between (S)- and (R)-propafenone, as a model substance to study the mechanism underlying that interaction in human liver microsomes. We used the cytochrome P450IID6-mediated 5-hydroxylation of propafenone as a model pathway, because this metabolic step constitutes the major route of biotransformation of propafenone. The Michaelis-Menten kinetics for 5-hydroxylation were determined after incubation of (R)- and (S)-propafenone and a pseudoracemate consisting of (S)-[2H4]propafenone and (R)-propafenone. Inhibition experiments were performed using (S)-[2H4]propafenone as an inhibitor of the 5-hydroxylation of (R)-propafenone, and vice versa. The kinetic model of mixed alternative substrates was used to simulate inhibition experiments. Experimental data were compared with those predicted by this model. We observed a substantial stereoselectivity after incubation of the individual enantiomers [(S)-propafenone: Vmax, 10.2 pmol/micrograms/hr, and Km, 5.3 microM; (R)-propafenone: Vmax, 5.5 pmol/micrograms/hr, and Km, 3.0 microM]. In contrast, no substrate stereoselectivity was observed after incubation of the pseudoracemate [3.1 pmol/micrograms/hr for (S)-[2H4]propafenone and 3.3 pmol/micrograms/hr for (R)-propafenone]. Application of the model revealed Ki values of 2.9 and 5.2 microM for the inhibition of 5-hydroxylation of (S)-[2H4]-propafenone by (R)-propafenone and for inhibition of 5-hydroxylation of (R)-propafenone by (S)-[2H4]-propafenone, respectively. The predicted and the experimental data were in good agreement, and both indicated the mode of inhibition to be competitive. In conclusion, the enantiomers of propafenone interact with respect to 5-hydroxylation, with (R)-propafenone being a more potent inhibitor than the S-enantiomer with respect to cytochrome P450IID6-mediated 5-hydroxylation. Because beta-blocking properties of propafenone reside in the S-enantiomer, inhibition of metabolism of this enantiomer by (R)-propafenone may have therapeutic consequences.     

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.     

Effect of selective monoamine oxidase inhibition by clorgyline and deprenyl on the norepinephrine receptor-coupled adenylate cyclase system in rat cortex

The consequences of selective monoamine oxidase (MAO) inhibition on the norepinephrine(NE)-sensitive adenylate cyclase system were determined in slices of rat cerebral cortex. The chronic administration of clorgyline, which selectively inhibited the activity of MAO-A, caused a significant decrease in the responsiveness of the noradrenergic cyclic AMP-generating system. The noradrenergic subsensitivity was accompanied by a significant decrease in the density of beta-adrenoceptors, as measured by 3H-dihydroalprenolol (DHA) binding, without altering the Kd value. However, selective inhibition of MAO-B by deprenyl did not alter the sensitivity of the cyclic AMP-generating system to NE or the specific DHA binding. The basal levels of cyclic AMP in the cortex were unaltered by the drugs. Since inhibition of MAO-A, but not MAO-B, increases the availability of NE, the results support the hypothesis that a persistent NE-receptor interaction is one of the prerequisites for the in vivo densitization of the NE-sensitive adenylate cyclase and the concomitant down-regulation of the number of beta-adrenoceptors in brain.     

Synthesis and monoamine oxidase inhibitory activity of new pyridazine-, pyrimidine- and 1,2,4-triazine-containing tricyclic derivatives

A number of condensed azines, mostly belonging to the families of indeno-fused pyridazines (1), pyrimidines (2, 3), and 1,2,4-triazines (4, 5), were synthesized and evaluated in vitro as monoamine oxidase (MAO) A and B inhibitors. Most of them showed higher inhibition potency toward MAO-B, the most effective one being 3-(3-nitrophenyl)-9H-indeno[1,2-e] [1,2,4]triazin-9-one (4c), which displayed an IC50 value of 80 nM and proved to be 10-fold more potent than its [2,1-e] fusion isomer 5. Replacing the 3-phenyl group of the known indeno[1,2-c]pyridazin-5-one MAO-B inhibitors with a flexible phenoxymethyl group enhanced the inhibitory potency. The inhibition data highlighted the importance of the aza-heterocyclic scaffold in affecting the MAO isoform selectivity. The 3-phenyl derivatives with type 1, 4, and 5 scaffolds were inhibitors of MAO-B with little or no MAO-A effect, whereas 2- or 3-phenyl derivatives of type 2 and 3 pyrimidine-containing fusion isomers inhibited both isoenzymes with a structure-dependent preference toward MAO-A.