Frequent administration of levodopa/carbidopa microtablets vs levodopa/carbidopa/entacapone in healthy volunteers.

OBJECTIVES: An oral dispersible microtablet formulation of levodopa/carbidopa 5/1.25 mg (LC-5) was developed for individualized repeated dosing. The aim was to compare pharmacokinetic profiles of LC-5 and levodopa/carbidopa/entacapone (LCE). MATERIALS AND METHODS: A randomized, crossover study was carried out in 11 healthy subjects. Plasma concentrations of levodopa, carbidopa and 3-O-methyldopa were determined after intake of 300 mg levodopa during the day, either as three intakes of 100/25/200 mg LCE or as a morning dose of 75/18.25 mg followed by five repeated doses of 45/11.25 mg LC-5. RESULTS: Repeated dosing (2.4-hourly) with LC-5 microtablets compared to LCE (6-hourly) avoided long periods with low plasma levodopa levels. Time to maximum plasma concentrations was significantly shorter for LC-5. LC-5 showed lower fluctuation index (FI) in plasma compared to LCE (ANOVA P = 0.0028). FI for dose 2-5 was on average 1.26 for levodopa in LC-5, and 2.23 for dose 1-2 of LCE. The ratio between the two mean FI:s is 0.565; that is, LC-5 gave nearly half the FI as compared to LCE. CONCLUSIONS: Fractionation of levodopa with LC-5 into small, frequent administrations as compared to standard administrations of LCE decreased the FI in plasma for both levodopa and carbidopa by nearly half.

Effects of the nitrone radical scavengers PBN and S-PBN on in vivo trapping of reactive oxygen species after traumatic brain injury in rats.

In previous studies, the authors showed that the nitrone radical scavenger alpha-phenyl-N- tert -butyl nitrone (PBN) and its sulfo-derivative, 2-sulfo-phenyl-N- tert -butyl nitrone (S-PBN), attenuated cognitive disturbance and reduced tissue damage after traumatic brain injury (TBI) in rats. In the current study, the production of reactive oxygen species (ROS) after TBI was monitored with microdialysis and the 4-hydroxybenzoic acid (4-HBA) trapping method. A single dose of PBN (30 mg/kg) or an equimolar dose of S-PBN (47 mg/kg) was administered intravenously 30 minutes before a controlled cortical contusion injury in rats. Plasma and brain tissue drug concentrations were analyzed at the end of the microdialysis experiment (3 hours after injury) and, in a separate experiment with S-PBN, at 30 and 60 minutes after injury. Traumatic brain injury caused a significant increase in ROS formation that lasted for 60 minutes after the injury as evidenced by increased 3,4-dihydroxybenzoic acid (3,4-DHBA) concentrations in the dialysate. PBN and S-PBN equally and significantly attenuated the posttraumatic increase in 3,4-DHBA formation. High PBN concentrations were found bilaterally in brain tissue up to 3 hours after injury. In contrast, S-PBN was rapidly cleared from the circulation and was not detectable in brain at 30 minutes after injury or at any later time point. The results suggest that scavenging of ROS after TBI may contribute to the neuroprotective properties observed with nitrone spin-trapping agents. S-PBN, which remained undetectable even in traumatized brain tissue, reduced ROS production to the same extent as PBN that readily crossed the blood-brain barrier. This finding supports an important role for ROS production at the blood-endothelial interface in TBI.

Derivatives of 2-(dipropylamino)tetralin: effect of the C8-substituent on the interaction with 5-HT1A receptors.

A series of 2-(dipropylamino)tetralin derivatives in which the C8 substituent is varied has been prepared and evaluated pharmacologically to explore the importance of the C8 substituent in the interaction of 2-aminotetralin-based ligands with serotonin (5-HT1A) receptors. Enantiopure derivatives were prepared by facile palladium-catalyzed reactions of the triflates of the enantiomers of 8-hydroxy-2-(dipropylamino)tetralin (8-OH-DPAT, 1). The affinity of the compounds for the 5-HT1A receptors was evaluated by competition experiments with [3H]-8-OH-DPAT in rat hippocampal and cortical tissue. In addition, the compounds were evaluated for central 5-HT and dopamine receptor stimulating activity in vivo by use of biochemical and behavioral assays in rats. With the exception of the carboxy-substituted derivative which is devoid of 5-HT1A receptor affinity, the compounds have moderate to high affinities (K(i) values range from 0.7 to 130 nM) for 5-HT1A receptors. Surprisingly, several of the derivatives do not produce any apparent effects in vivo although they have fairly high 5-HT1A receptor affinities. However, the methoxycarbonyl- and acetyl-substituted derivatives are potent 5-HT1A receptor agonists in vivo and exhibit in vitro affinities in the same range as the enantiomers of 1.

Derivatives of cis-2-amino-8-hydroxy-1-methyltetralin: mixed 5-HT1A-receptor agonists and dopamine D2-receptor antagonists.

(1S,2R)-8-Hydroxy-1-methyl-2-(dipropylamino)tetralin [(1S,2R)-3] has been previously characterized as a selective and potent but partial 5-HT1A-receptor agonist. In the present study, we have prepared derivatives of (1S,2R)- and (1R,2S)-3 in which various C8-substituents have been introduced. In addition, the enantiomers of the N-isopropyl-N-n-propylamino derivative of 3 were prepared. The new derivatives were tested in vivo by use of behavioral and biochemical tests in rats. In addition, the affinity of the compounds was studied by competition experiments with [3H]-8-OH-DPAT in rat brain tissue. The only new derivative which behaved like a selective 5-HT1A-receptor agonist was the C8-carboxamide derivative (1S,2R)-13. The other active derivatives, including (1S,2R)-3, have more complicated pharmacological profiles and may be best characterized as mixed 5-HT1A-receptor agonists/dopamine D2-receptor antagonists.

Increased total activity in the rat after L-tryptophan plus the monoamine oxidase-A inhibitor amiflamine but not after L-tryptophan plus clorgyline.

The effect of pretreatment with either saline or the monoamine oxidase-A inhibitors clorgyline and amiflamine upon the total activity, locomotion and rearing behaviour of the rat induced by various doses of the monoamine precursor L-tryptophan was studied by use of automated activity boxes. Amiflamine (2.5 and 5.0 mg kg-1, i.p.) increased in a dose-dependent manner total activity and to a lesser extent, locomotion when given 60 min before L-tryptophan (100 mg kg-1, i.p.). The increased activity was seen after amiflamine plus either 25 or 75 mg kg-1 L-tryptophan. Rearing behaviour was not affected. Analysis of 5-hydroxytryptamine (5-HT) and its deaminated metabolite 5-hydroxyindoleacetic acid (5-HIAA) by high performance liquid chromatography with electrochemical detection indicated that in both frontal cortex and hypothalamus, amiflamine (at both doses) increased 5-HT and reduced 5-HIAA concentrations. Combination of amiflamine with L-tryptophan (100 mg kg-1, i.p.) resulted in a higher 5-HT concentration being found than after amiflamine alone. L-Tryptophan treatment alone did not change 5-HT concentrations but increased 5-HIAA concentrations. Clorgyline, at a dose of either 1 or 5 mg kg-1 i.p. plus L-tryptophan (25 or 100 mg kg-1, i.p.) did not increase total activity, locomotion or behaviour. A number of possible explanations for the differences in the behavioural effects of clorgyline and amiflamine when given with L-tryptophan are discussed. It is concluded that in addition to monoamine oxidase-A inhibition, other pharmacological effects of the drugs, such as 5-HT release (amiflamine) and inhibition of tryptophan hydroxylation (clorgyline) may be of importance in determining the magnitude of the increase in activity when the compounds are given together with L-tryptophan.

Pharmacogenomics to predict drug response.

From theory to proof-of-concept, pharmacogenomics promises to improve future general healthcare in a number of ways. By identifying individuals who will respond to a particular drug treatment compared to those who have a low probability of response, pharmacogenomic test development hopes to aid the physician in prescribing the optimal medication for each patient. This approach promises faster relief from symptoms, a lowering of side effect risks and a reduction in healthcare costs. Pharmacogenomic tests used by the pharmaceutical companies themselves can be used to help identify suitable subjects for clinical trials, aid in interpretation of clinical trial results, find new markets for current products and speed up the development of new treatments and therapies. This type of approach should also see fewer compounds failing during later phases of development. The questions we are faced with as we enter the new millennium, however, are if and when the promises of pharmacogenomnics in improving healthcare will be fulfilled. Currently, there are only a handful of pharmacogenomic tests and associated products which are commercially available and it remains to be seen what impact these will have on the market and on healthcare in general.

Monitoring of reactive oxygen species production after traumatic brain injury in rats with microdialysis and the 4-hydroxybenzoic acid trapping method.

The detection of reactive oxygen species (ROS) after traumatic brain injury (TBI) is based on indirect methods due to the high reactivity and short half-life of ROS in biological tissue. The commonly used salicylate trapping method has several disadvantages making it unsuitable for human use. We have evaluated 4-hydroxybenzoic acid (4-HBA) together with microdialysis (MD) in the rat as an alternative method. 4-HBA forms one stable adduct, 3,4-dihydroxybenzoic acid (3,4-DHBA), when reacting with ROS and has not previously been used together with MD after TBI. Twenty-seven rats were used for the assessment of 3,4-DHBA production as an indicator of ROS formation in a controlled contusion injury model using intracerebral MD with 3 mM 4-HBA in the perfusate. For comparison, salicylate trapping was used in eight rats. TBI caused a 250% increase of 3,4-DHBA that peaked at 30 min after injury in severely injured rats and remained significantly elevated as compared to baseline for 90 min after trauma. The mild injury level caused a 100% increase in 3,4-DHBA formation at 30 min after the injury. When the MD probe was placed in the perimeter of the injury site, no significant increase in ROS formation occurred. Salicylate trapping showed a similar increase in adduct formation after severe injury. In addition, high cortical concentrations of 4-HBA and salicylate were found. It is concluded that microdialysis with 4-HBA as a trapping agent appears to be a useful method for ROS detection in the rat with a potential clinical utility.

Tolerance to alpha-methyl-p-tyrosine in rats: studies on the antagonism of amphetamine-induced motor activity and excitatory behaviour.

Rats were treated chronically with alpha-methyl-p-tyrosine methyl-ester HCl (alpha-MT) twice daily for 0--14 days. At 1 h after the (last) alpha-MT injection, d-amphetamine sulphate was given and motor activity was measured in an ANIMEX activity meter for 4 h. Amphetamine-induced excitatory and stereotyped behaviour was scored according to a rating scale in a separate experiment. A single dose of alpha-MT markedly reduced the activity response after amphetamine. After 1--3 days of alpha-MT treatment, tolerance to its amphetamine-antagonistic affect started to develop, reaching a maximal degree after 7--14 days. The pattern of the amphetamine response, monophasic in control rats, became biphasic in the alpha-MT tolerant rats with an early (at 0--1 h) and a late (2--4 h) peak of motor activity. The late peak appeared within 3 days, while the early peak appeared after 7 days of alpha-MT treatment. The results on amphetamine-induced excitatory and stereotyped behaviour in essence agreed with the motor-activity data. It is concluded that tolerance to the amphetamine-antagonistic action of alpha-MT is not complete. Its rate of development varies in a complex pattern, indicating the presence of more than one mechanism of tolerance.

Effects of d-amphetamine and methylphenidate on hyperactivity produced by neonatal 6-hydroxydopamine treatment.

Neonatal intracisternal administration of 6-hydroxydopamine (6-OHDA, 50 micrograms on day 1 after birth) caused a marked hyperactivity when the rats were tested as adults. These rats also showed severe DA depletions in striatum and nucleus accumbens. Pretreatment with the noradrenaline (NA) uptake inhibitor desipramine provided protection against NA depletion in frontal cortex and nucleus accumbens. Pretreatment with DA uptake inhibitors, amfolenic acid or GBR 12909, before 6-OHDA, provided full protection against DA depletion but produced marked NA depletion in frontal cortex. These rats did not demonstrate any degree of hyperactivity. Low doses of d-amphetamine (0.25 mg/kg SC) or methylphenidate (1 mg/kg SC) reversed the hyperactivity in DA-depleted rats but increased motor activity in vehicle-treated and NA-depleted rats. Higher doses of d-amphetamine (1 mg/kg) or methylphenidate (4 mg/kg) produced potentiated levels of locomotion but attenuated levels of rearing in DA-depleted animals. The results further suggest the utility of the neonatal DA lesion in rats as a potential animal model for derivation of therapeutic agents that may be efficacious in the treatment of the hyperkinetic syndrome.

Central noradrenaline depletion antagonizes aspects of d-amphetamine-induced hyperactivity in the rat.

The effects of noradrenaline (NA) depletion upon amphetamine-induced hyperactivity were examined in five experiments. Central NA depletion via either systemic DSP4 or neonatal 6-OHDA antagonised the amphetamine-induced (2 mg/kg SC) increase in rearing behaviour, whereas lesions of the dorsal noradrenergic bundle using 6-hydroxydopamine antagonised the increase in locomotor activity. Peripheral NA depletion following systemic 6-hydroxydopamine to adult rats did not cause any changes in motor activity after acute amphetamine administration. Desipramine, the selective NA uptake inhibitor, blocked the effects of DSP4 upon amphetamine-induced rearing. NA depletion antagonised hyperactivity produced by the 2 mg/kg dose of amphetamine, but not the hyperactivity (rearing or locomotion) effects of amphetamine at 1, 4 or 8 mg/kg.

(S)-UH-301 antagonizes (R)-8-OH-DPAT-induced cardiovascular effects in the rat.

The 5-HT1A-receptor antagonist (S)-UH-301 (S)-5-fluoro-8-hydroxy-2- (dipropylamino)tetralin) completely antagonized the hypotension and bradycardia induced by (R) = 8-OH DPAT [R)-8-hydroxy-2- (dipropylamino)tetralin) in conscious rats. (S)-UH-301 alone induced a weak hypertension, which might be due to its 5-HT1A-receptor antagonistic properties. (R)-UH-301 induced effects similar to those of (R)-8-OH DPAT, i.e., a short initial phase of hypertension followed by a long-lasting hypotension and bradycardia. Thus, (R)-UH-301 behaves as a 5-HT1A-receptor agonist and (S)-UH-301 as a 5-HT1A-receptor antagonist, abolishing the effects induced by (R)-8-OH DPAT.

(R)- and (S)-8-acetyl-2-(dipropylamino)tetralin (LY-41): two novel 5-HT1A receptor agonists.

Interactions between central 5-HT1A receptors and the enantiomers of LY-41, a 2-aminotetralin derivative related to 8-OH-DPAT (8-hydroxy-2-(dipropylamino)tetralin), were studied. Both enantiomers of LY-41 behaved as potent 5-HT1A receptor agonists in rats, inducing the 5-HT behavioural syndrome, decreasing body temperature and inhibiting the cage-leaving response. The behavioural syndrome and the hypothermia were antagonized by the 5-HT1A receptor antagonist, (S)-UH-301. The LY-41 enantiomers also reduced brain 5-HTP accumulation in rats treated with a decarboxylase inhibitor. The pharmacology of the enantiomers of LY-41 appeared similar to that of 8-OH-DPAT. However, it is noteworthy that the stereoselective interaction of 5-HT1A receptors with LY-41 was opposite to that of 8-OH-DPAT. Thus, (R)-8-OH-DPAT was more potent than (S)-8-OH-DPAT, whereas (S)-LY-41 appeared to be more potent than (R)-LY-41.

The pharmacological assessment of a new, potent oxotremorine analogue in mice and rats.

Some of the central and peripheral effects of oxotremorine (OT) and its azetidine analogue, N-[4-azetidinyl)-2-butynyl]-2-pyrrolidone (BM 120), were compared in mice and rats. BM 120 was found to be about twice as potent as OT and to have a somewhat longer duration of action. All its effects were antagonized by pretreatment with atropine sulphate. BM 120 acted as powerful muscarinic agonist on the isolated guinea pig ileum.

Differential serotoninergic and dopaminergic activities of the (R)- and the (S)-enantiomers of 2-(di-n-propylamino)tetralin.

Racemic 2-(di-n-propylamino)tetralin ((R,S)-DPAT), which lacks phenolic or other aromatic substituents, induces both dopaminergic (sniffing, licking and gnawing) and serotoninergic (forepaw treading and flat body posture) behavioural responses. The present study shows that s.c. administration of (R)-DPAT induces typical 5-HT1A receptor agonist behaviours. These effects are blocked by the 5-HT1A receptor antagonist (S)-5-fluoro-8-hydroxy-2-(di-n-propylamino)tetralin ((S)-UH-301). Administration of (S)-DPAT induces dopaminergic behaviours, which are fully antagonised by raclopride, a dopamine D2 receptor antagonist. Both enantiomers induce hypothermia, (R)-DPAT being antagonised by (S)-UH-301, whereas (S)-DPAT is antagonised by raclopride. The accumulation of 5-hydroxytryptophan and DOPA (3,4-dihydroxyphenylalanine) after decarboxylase inhibition that reflects presynaptic actions on 5-HT (5-hydroxytryptamine, serotonin) and dopamine neurons, respectively, are inhibited by both enantiomers of DPAT. (R)-DPAT is more potent than (S)-DPAT as an inhibitor of 5-hydroxytryptophan accumulation whereas (S)-DPAT is more potent than (R)-DPAT as an inhibitor of DOPA accumulation. Thus, in functional tests of postsynaptic actions (R)-DPAT behaves as a 5-HT1A receptor agonist and (S)-DPAT as a dopamine D2 receptor agonist. Presynaptically, (R)-DPAT shows selectivity for 5-HT1A receptors and (S)-DPAT for dopamine D2 receptors. Receptor binding studies, utilizing [3H]8-hydroxy-2-(di-n-propylamino)tetralin and [3H]quinpirole as radioligands for 5-HT1A and dopamine D2 receptors, respectively, showed (R)-DPAT to have a 3-fold higher affinity than (S)-DPAT for 5-HT1A receptors, whereas (S)-DPAT had a 6-fold higher affinity than (R)-DPAT for dopamine D2 receptors. Thus, the results from receptor binding studies support the conclusion that (R)- and (S)-DPAT are agonists showing selectivity for 5-HT1A and dopamine D2 receptors, respectively. Taken together, these findings may explain previous controversies with regard to the pharmacology of racemic DPAT and re-emphasise the necessity to study pure enantiomers of chiral compounds.

Pharmacokinetic and pharmacodynamic studies of (R)-8-hydroxy-2-(di-n-propylamino)tetralin in the rat.

Racemic 8-OH-DPAT, (R,S)-8-hydroxy-2-(di-n-propylamino)tetralin, has become the prototype 5-HT1A receptor agonist. The enantiomers of 8-OH-DPAT have similar affinities to the 5-HT1A receptor, but the (R)-enantiomer is a full agonist, whereas the (S)-enantiomer is a partial agonist. This communication describes the dose- and time-response relationships of behavioural (5-HT behavioural syndrome, cage-leaving response), physiological (body temperature) and biochemical (5-HT turnover, 5-hydroxytryptophan accumulation) effects of (R)-8-OH-DPAT in rats. A high-performance liquid chromatography (HPLC)-UV method for determination of plasma and brain concentrations of (R)-8-OH-DPAT was developed, permitting studies of the pharmacokinetics of the drug. The concentrations of 8-OH-DPAT in brain were several fold higher than in plasma, and there were large variations in (R)-8-OH-DPAT concentrations between brain regions (highest in the hippocampus). (R)-8-OH-DPAT peaked in plasma at 5 min and in brain at 15 min after subcutaneous administration. The 5-HT1A behavioural syndrome peaked within 5 min after administration and disappeared after 30 min, when brain concentrations were still high. The hypothermic and biochemical responses developed gradually and were maximal at 45-60 min post injection, when both plasma and brain concentrations were declining. Thus, there was not a simple relationship between the kinetics and the dynamics of (R)-8-OH-DPAT. These results prompt further studies on the pharmacokinetics of 8-OH-DPAT within the central nervous system.

Differential tissue distribution of the enantiomers of racemic pindolol in the rat.

Racemic pindolol, a beta-adrenoceptor and a 5-HT1A/1B receptor antagonist, has been reported to augment and accelerate the clinical efficacy of antidepressants. The (S)-enantiomer is more potent than the (R)-enantiomer both at the beta-adrenergic and at the 5-HT1A/1B receptors. A chiral HPLC column was used for determination of tissue concentrations of the enantiomers of pindolol at 90 min after 8 mg/kg s.c. of the racemate. The (S)/(R) ratio was found to vary between tissues from 1.74 in brain to 0.82 in plasma. The present findings may be important for understanding the pharmacodynamic actions of racemic pindolol.

Effects of (R)-8-OH-DPAT and the enantiomers of UH-301 on motor activities in the rat: antagonism of (R)-8-OH-DPAT-induced effects.

The effects of the enantiomers of 5-fluoro-8-hydroxy-2-(dipropylamino)tetralin, UH-301 and the potent 5-HT1A-receptor agonist (R)-8-hydroxy-2-(dipropylamino)tetralin, (R)-8-OH-DPAT, on locomotion, rearing and total activity were studied in rats. The experiments were performed as tests either of exploratory activity in non-habituated rats or of motor activity of rats habituated to the environment for 2 h before drug injection. (R)-8-OH-DPAT increased locomotion and total activity and decreased rearing in both conditions. (R)-UH-301 increased locomotion and slightly also total activity in habituated rats and decreased rearing in both conditions. (S)-UH-301 decreased locomotion and rearing in both conditions but only in doses of 10 mumol/kg and above. Lower doses of (S)-UH-301 (10 mumol/kg) antagonized (R)-8-OH-DPAT-induced increases of locomotion and total activity. As (S)-UH-301 decreased rearing, per se, it was not able to antagonize the (R)-8-OH-DPAT induced decrease. These results further support previous data that (S)-UH-301 is a 5-HT1A antagonist while (R)-UH-301 is a 5-HT1A agonist.

Ethanol-induced hypothermia and biogenic amine metabolites.

Ethanol is known to cause hypothermia. The rectal temperature of rats receiving ethanol, 4 g/kg i.p., at an ambient temperature of 23 degrees C decreased by 2 degrees C. This body temperature decrease could be prevented by keeping the animals at an ambient temperature of 34 degrees C. Irrespective of the body temperature it was found that the concentration of the major metabolites of dopamine and serotonin in brain tissue was significantly increased. Thus, the change in brain monoamine metabolite levels in rats after administration of ethanol are not due to ethanol-induced hypothermia.

Inhibition of the in vivo biosynthesis and changes of catecholamine levels in rat brain after alpha-methyl-p-tyrosine; time- and dose-response relationships.

Male Sprague-Dawley rats were given 0.407 mmoles/kg of D,L-alpha-methyl-p-tyrosine methylester HCl (H44/68; alpha-MT) at eleven time-points between 0--24 h, or 8 doses between 0.013--1.628 mmoles/kg of the drug at 1 h before i.v. injection of 160 micronCi tyrosine-2,6-3H. The rats were killed 15 min after tyrosine-3H and brain alpha-MT, tyrosine and catecholamines (endogenous and labelled), and plasma alpha-MT and tyrosine (--3H) were chromatographically isolated before being assayed spectrophotofluorimetrically (endogenous) or by liquid scintillation methods (labelled compounds). A delayed penetration of alpha-MT from plasma into brain, different elimination rates of alpha-MT in plasma and brain, and decreasing brain/plasma drug concentration on increasing alpha-MT dosages, indicated, that alpha-MT in brain and plasma belong to different pharmacokinetic compartments. The endogenous levels of catecholamines in the time-response experiments, declined to a minimum 4 h after alpha-MT administration, where the dopamine level was 38% and the noradrenaline level 51% of the saline controls. Kinetic data of the catecholamine elimination is given. In the dose-response experiment the decrease in the endogenous catecholamine levels was dose-related up to 0.407 mmoles/kg of alpha-MT, with no further decline on higher doses. The maximal inhibition of brain catecholamine synthesis occurred within 30 min after alpha-MT administration and the inhibition correlated better with the brain than with plasma alpha-MT content. The inhibition was dose-related with a maximal synthesis inhibition of 95% for dopamine and 80% for noradrenaline at the highest dose of alpha-MT. The duration of synthesis inhibition and storage depletion were shorter for noradrenaline (12 h) than for dopamine (16 h). Further, the ED50 for synthesis inhibition of dopamine (0.057 mmoles/kg) was half of the ED50 for synthesis inhibition of noradrenaline (0.117 mmoles/kg). This might suggest different sensitivities towards alpha-MT or different availabilities of alpha-MT in the two neuron populations. At the three highest doses of alpha-MT there were signs of interference with the uptake process for tyrosine from plasma into the brain. This was indicated by increased plasma levels and decreased brain levels of tyrosine (--3H).

The relationship between amphetamine antagonism and depletion of brain catecholamines by alpha-methyl-p-tyrosine in rats.

The time-course and the dose-response relationship for the antagonistic effect of alpha-methyl-p-tyrosine methyl ester HCl H 44/68 (alpha-MT) on d-amphetamine (10.6 mumoles/kg) induced increase in motor activity was studied. The effect of amphetamine was gradually reduced from 30--60 min to a minimum at 1--4 h after the administration of 0.407 mmoles/kg of alpha-MT. From (4--) 8 h the amphetamine response started to reappear and the original response was restored completely at 16 h after alpha-MT. The dose-response curve showed, that between 0.051--0.41 mmoles/kg of alpha-MT, given 1 h before amphetamine, there was a gradual reduction of the amphetamine response; doses above 0.41 mmoles/kg did not cause any further effect. The antiamphetamine action of alpha-MT was compared with its time- and dose-dependent effects of inhibition of synthesis and reduction of stores of brain catecholamines. It was found, that the antiamphetamine action was more closely correlated with the reduction of the levels of brain dopamine, than with the brain noradrenaline levels. Further, the inhibition of catecholamine synthesis per se did not appear to be a sufficient condition for alpha-MT induced antagonism of amphetamine. These findings support the view that amphetamine is dependent on a substantial portion of the brain pool of dopamine and possibly noradrenaline rather than on very small, newly synthesized pools of these neurotransmitters.