Kazuya Kamijo

Substrate Selectivity of Type A and Type B Monoamine Oxidase in Rat Brain

Abstract: Use of the irreversible inhibitors clorgyline and deprenyl showed that rat brain mitochondria contain type A and type B monoamine oxidase (MAO). Tyramine is a substrate for both types of MAO, whereas serotonin is a preferential substrate for type A MAO. In contrast to MAO in other tissues, type A MAO in brain tissue oxidizes β‐phenylethylamine (PEA) at high concentrations (0.5 and 1.0 mM). The proportions of type A and type B MAO activities in the mitochondria estimated from the double‐sigmoidal inhibition curves of tyramine oxidation were about 70:30 irrespective of the concentration of tyramine. With PEA as substrate, the ratios of type A to type B activities were found to increase from low values at low concentrations to about 1 at 0.5‐1.0 mM‐PEA, and even higher at further increased concentrations of PEA. At very low (0.01 mM) and high (10.0 mM) concentrations of PEA, single‐sigmoidal curves were obtained; with the high PEA concentration the activity was highly sensitive to clorgyline, whereas with the low concentration it was highly sensitive to deprenyl. In deprenyl‐pretreated mitochondrial preparations, all the remaining activity towards 0.5‐1.0 mM‐PEA was shown to be highly sensitive to clorgyline, demonstrating that this activity was indeed due to oxidation by type A MAO. The opposite result was obtained with deprenyl as inhibitor of clorgyline‐pretreated preparations, demonstrating that PEA at this concentration was also oxidized by type B MAO in rat brain mitochondria. The K3 values of type A and type B MAO for PEA were significantly different. On Lineweaver‐Burk analysis, plots with PEA as substrate for type A MAO in a deprenyl‐treated preparation were linear over a wide concentration range, whereas those for type B MAO in a clorgyline‐treated preparation were not linear, but showed substrate inhibition at higher concentrations of the substrate. It is concluded from the present findings that the effect of the substrate concentration must be considered in studies on the characteristics of multiple forms of MAO in various organs and species.

Multiple catalytic sites of rat brain mitochondrial monoamine oxidase.

Abstract We compared the inhibitory and catalytic effects of various monoamines on forms A and B of monoamine oxidase (MAO) on mitochondrial preparations from rat brain in mixed substrate experiments. MAO activity was determined by a radioisotopic assay. MAO showed lower K m values for tryptamine and β-phenylethylamine than for tyramine and serotonin. The K m values of the untreated preparation for tyramine, tryptamine, and β-phenylethylamine obtained were the same as those of the form B enzyme and the K m value for serotonin was the same as that of the form A enzyme. Tyramine and tryptamine were competitive inhibitors of serotonin oxidation and β-phenylethylamine did not bind with form A enzyme or inhibit the oxidation of serotonin, while tyramine and tryptamine were competitive inhibitors of β-phenylethylamine oxidation. Although serotonin was not oxidized by form B enzyme, serotonin was a competitive inhibitor of β-phenylethylamine oxidation. It is suggested that rat brain mitochondrial MAO is characterized by two kinds of binding sites.

Monoamine oxidase in frog liver and brain

Abstract 1. 1. Studies with clorgyline and deprynyl as inhibitors showed that there are two types of mitochondrial flavoprotein monoamine oxidase (MAO) in frog brain and liver that resemble type A and type B MAO in many mammalian organs. 2. 2. The enzymic reactions occured by a double-displacement (ping-pong) mechanism. 3. 3. In frog liver, there is also another type of mitochondrial MAO, which contains copper and pyridoxal phosphate as cofactor, but its activity is very low with benzylamine as substrate.

Pharmacokinetics of Amikacin and Cephalothin in Bedridden Elderly Patients

Abstract: The pharmacokinetics of amikacin (5.5 mg/kg intramuscularly) and cephalothin (1000 mg/body intravenously) in bedridden elderly patients were studied in comparison with those in healthy volunteers. The eliminations of amikacin and cephalothin from the plasma followed the course of a one‐compartment open model. For amikacin, five healthy volunteers, elimination rate constant Kel was 0.396 hr−1 biologic half‐life t½ was 1.80 hour, volume of distribution Vd was 0.201 l./kg; in five bedridden elderly patients, Kel was 0.208 hr−1, t½ was 3.55 hours, Vd was 0.376 l./kg. Cumulative renal excretion of amikacin in 8 hours was 44 per cent of the total dose in bedridden elderly patients and 69 per cent in healthy volunteers. For cephalothin, in seven healthy volunteers, Kel was 0.0353 min−1, t½ was 19.7 min, Vd was 0.176 l./kg; in four bedridden elderly patients, Kel was 0.0127 min−1, t½ was 56.4 min, Vd was 0.283 l./kg. Cumulative renal excretion of cephalothin reached a plateau by 4 hours of 40.8 per cent of the total dose in bedridden elderly patients and of 56.7 per cent in healthy volunteers. These results suggest that in bedridden elderly patients decreased renal excretion of amikacin and cephalothin is related to decreased renal function and an increased Vd.

NONEXISTENCE OF A TYPE C MONOAMINE OXIDASE IN RAT BRAIN

Abstract— The possible existence of type C MAO, distinct from type A and type B, in circumventricular structures of rat brain was examined by histological studies on the inhibitory effects of clorgyline. a preferential type A MAO inhibitor and deprenyl, a preferential type B inhibitor, on enzyme. Brain slices were preincubated with the inhibitors and then incubated with 5‐HT, the substrate for type A MAO, and stained for MAO activity. Deposits of the product formazan were detected in circumventricular structures of slices of brain preincubated with clorgyline and deprenyl at concentrations of 10‐7–10‐4m at room temperature for 5 min. When the slices were preincubated with either of these inhibitors at room temperature for 60 min, strong activity was observed in this region, whereas when they were preincubated with either 10‐5m‐clorgyline or 10‐5m‐deprenyl for 20 and 30 min at 37°C, no MAO activity was seen in any region of the brain. Thus, at the higher preincubation temperature, lower concentrations of each inhibitor and a shorter preincubation period were required for inhibition of the enzyme. Preincubation for 60 min at 37°C with a combination of 10‐7m‐clorgyline and 10‐8m‐deprenyl did not inhibit the enzyme in the circumventricular region completely, but at the same temperature, concentrations of 10‐7m of both inhibitors inhibited the enzyme completely in 10min, Thus the effects of the inhibitors are synergistic. These results indicate that the inhibitory effects of the two inhibitors on the enzyme in circumventricular structures of the brain is time‐ and temperature‐dependent. Moreover, the activity seems to be sensitive to deprenyl even when 5‐HT is used as substrate. The results do not support the idea of the existence of type C MAO, distinct from type A and type B MAO.

Membrane lipid environment of carp brain and liver mitochondrial monoamine oxidase

Delipidation of carp liver mitochondria by treatment with methyl ethyl ketone (MEK) or Triton X-100 and then perchlorate greatly reduced MAO activities. Treatment with only Triton X-100 resulted in less reduction in activity. The Km values of the remaining activities were similar regardless of these treatments. The sensitivities towards clorgyline and l-deprenyl of the remaining activity in the Triton X-100-treated residue and the phospholipase C-treated carp brain mitochondria were found to be unchanged, but those of the activity remaining in the MEK-treated residue were similarly decreased. No evidence was obtained suggesting conversion of carp MAO to either MAO-A or MAO-B by the modification of the mitochondrial lipid environment by the treatments employed.