Isami Kuruma

L-dopa-induced accumulation of 3-O-methyldopa in brain and heart

Abstract Administration of L- 14 C-dopa in rats causes a considerable rise of 14 C-3-O-methyldopa in brain and heart which is of longer duration than that of the other 14 C-catechol metabolites. Following repeated administration of small doses of L- 14 C-dopa, the 14 C-3-O-methyldopa accumulates, reaching a plateau after the third dose. Levels far in excess of those of the other 14 C-catechol metabolites are reached.

Comparative investigation of inhibitors of extracerebral dopa decarboxylase in man and rats

The decarboxylase inhibitor Ro 4–4602 [N1‐(dl‐seryl)‐N2‐(2,3,4‐trihydroxybenzyl)hydrazine] in doses of 3.4–34.0 μmol/kg (1–10mg/kg) progressively enhances the increase of dopa and 3‐O‐methyldopa and diminishes the rise of phenolcarboxylic acids in human plasma induced by administration of l‐dopa. A dose of 3.4 μmol/kg Ro 4–4602 is approximately equipotent to 17.0 μmol/kg MK 485 [β‐(3,4‐dihydroxyphenyl)‐α‐hydrazino‐α‐methyl‐dl‐propionie acid]. Ro 4–4602, combined with 2 mg/kg l‐dopa, causes a higher increase of catecholamines than 20 mg/kg l‐dopa alone in the striatum and probably in the hypothalamus of rat brain, whereas in the other brain areas the amine rise is equal after both treatments. MK 485 in doses equimolar to Ro 4–4602 has a less marked effect in all brain areas. Either inhibitor (10–2 μmol/kg) combined with 2 mg/kg l‐dopa causes markedly less increase of catecholamines than 20 mg/kg l‐dopa alone in the rat heart. Ro 4–4602, in small single doses, used at present in the treatment of Parkinsons syndrome, markedly inhibits the extracerebral decarboxylation of l‐dopa in man and rats and is more potent than MK 485.

Determination of endogenous GABA released from the cerebral cortex slices of the rat by high-performance liquid chromatography with a series-dual electrochemical detector

A new, simple, rapid and sensitive method for the determination of ?-aminobutylic acid (GABA) has been developed by high-performance liquid chromatography with electrochemical detection (LCEC). A new and unique technique for LCEC by using the reductive-oxidative mode of a dual electrochemical detector provided a simple and sensitive assay method for GABA. The standard curve was linear over a concentration range of 1-1000 ng. The detection limit for GABA was less than 0.5 ng. This new method was adapted to the assay of the transmitter released endogenously from the cerebral cortical slices of the rat. Endogenous GABA release evoked by high K(+) was reduced when superfusion was performed in the presence of 100 ?M forskolin.

The disposition of the tolcapone 3-O-methylated metabolite is affected by the route of administration in rats.

Abstract— Catechol‐O‐methyltransferase (COMT) catalyses the transfer of the methyl group from S‐adenyl‐l‐methionine (SAM) to one of the hydroxy groups of a catechol, usually the hydroxy group in position 3. COMT is present mainly in a soluble form (S‐COMT) in the cytosol, but a small fraction is bound to cell membranes (MB‐COMT). MB‐COMT has higher affinity for the catechol substrate than does S‐COMT by a factor of > 10, and high MB‐COMT activity is observed in the intestinal muscle layer. The present study investigates the effect of the administration route on the disposition of the tolcapone 3‐O‐methylated metabolite following intravenous and oral tolcapone administration in rats. Tolcapone is a substrate for COMT although the 3‐O‐methylated metabolite produced has no pharmacological actions. The 3‐O‐methylated metabolite was eliminated very slowly following oral administration of tolcapone, and its concentration approached a plateau level, which was in contrast to the situation following intravenous administration of tolcapone. It is thought that the oral dose of tolcapone receives a high exposure to MB‐COMT in the intestinal muscle layer during its absorption, and tolcapone seems to form a complex with MB‐COMT having a high affinity constant (i.e. a very low Ki). The fraction of the intravenous dose of tolcapone metabolized to the 3‐O‐methylated metabolite at 10 mg kg−1 was 2·6%, whereas those of the oral doses, which were corrected by the bioavailability, were 5·4% for 20 mg kg−1 and 2·7% for 40 mg kg−1.

Elevation of adenosine 3',5'-monophosphate in the perfusate of rat kidney after addition of dopamine.

Dopamine (10-4 M) and vasopressin (1 mU/ml) were found to increase the level of cyclic AMP in the perfusate of rat kidney. There were some differences in the mode of action of these two drugs. Firstly, the effect of dopamine, but not of vasopressin, was completely antagonized by spiroperidol. Secondly, the maximal response was attained within 1 min after dopamine perfusion, but 8 min after vasopressin perfusion. These results suggest that a specific dopamine receptor which acts to increase the concentration of cyclic AMP is located in the vascular tissue of rat kidney.

Estimation of kinetic parameters in the inactivation of an enzyme by a suicide substrate

A method was developed to estimate the extended Michaelis constant and maximum velocity of a suicide substrate from the time-course of remaining enzyme activity with the use of simulation data calculated from the representative kinetic model for a suicide substrate proposed by Walsh et al. (Walsh, C., Cromartie, T., Marcotte, P. and Spencer, R. (1978) Methods Enzymol. 53, 437-448). For this purpose an analytical equation for the time-course of remaining enzyme activity, based on the suicide kinetic model, was derived by the steady-state method reported by Tatsunami et al. (Tatsunami, S., Yago, N. and Hosoe, M. (1981) Biochim. Biophys. Acta 662, 226-235). The accuracy of this analytical solution was proved by comparing the result with the exact solution obtained by numerical computation. A method was also developed to estimate the most important factor for a suicide substrate, the partition ratio, from the time-course of remaining enzyme activity.

A possible intraneuronal site of action of thymoleptics.

The uptake of catecholamines (CAs) into crude mitochondria preparations (P2 fractions) and vesicle preparations from rat hypothalamus and striatum were compared in terms of the inhibition by thymoleptics and other drugs.Thymoleptics preferentially inhibited the uptake of CAs into hypothalamic P2 fractions, while ATPase inhibitors preferentially inhibited the uptake of dopamine into striatal P2 fractions. When the preparation obtained from rats pretreated with reserpine was used, the preferential inhibition of hypothalamic uptake by thymoleptics was entirely abolished. When P2 fractions from both regions were incubated with 10-6M 14C-imipramine, the intrasynaptosomal distribution of labeled imipramine revealed its affinity not only to the synaptosomal membrane, but also to the synaptic vesicles. Accumulated 3H-norepinephrine (NE) could be released by a hypoosomotic shock from striatal P2 fractions, but not from hypothalamic P2 fractions. The ATP-Mg2+-dependent uptake of 3H-NE into the synaptic vesicles from rat brain stem was inhibited by desipramine.These results indicate that the inhibition of CA uptake by thymoleptics in the hypothalamus is predominantly due to the inhibition at the synaptic vesicle, while in the striatum the uptake at the synaptosomal membrane is predominantly inhibited.

Lack of an Effect of Madopar on the Disposition of Tolcapone and its 3‐O‐Methylated Metabolite in Rats

The effect of Madopar (benserazide and l‐dopa, 1:4) on the disposition of the new selective inhibitor of catechol‐O‐methyltransferase, tolcapone, in rats was investigated. There was no statistically significant difference in the pharmacokinetic parameters of tolcapone in the presence or absence of Madopar except for a change in the mean residence time after oral administration. Thus, we rejected the hypothesis that the consumption of S‐adenyl‐l‐methionine by Madopar would change the disposition of tolcapone.

The behavior of remaining enzyme activity in a suicidal enzyme system

We derived an equation which describes the plot of the remaining enzyme activity versus ratio of initial concentration of suicide substrate to that of enzyme to obtain a partition ratio from the time-course of remaining enzyme activity. The simulation data calculated from the representative kinetic model for a suicide substrate were used to verify this equation, which approximated steady state kinetics. Although the time-dependent loss of enzyme activity is usually characterized by pseudo-first-order kinetics, the present results show that pseudo-first-order kinetics are followed only when the ratio of initial concentration of suicide substrate to that of enzyme is greater than the partition ratio. Our results also show that the present method can be used to obtain the partition ratio of a suicide substrate from the time-course of the remaining enzyme activity when the suicide substrate is given an arbitrary concentration of one, where the ratio of initial concentration of suicide substrate to that of enzyme is less than the partition ratio. The theoretically verified equation was also checked against reported experimental data for a microsomal enzyme system.