Interstitial 3-Methoxytyramine Reflects Striatal Dopamine Release: An In Vivo Microdialysis Study

Previous ex vivo studies have provided indirect evidence that the dopamine (DA) metabolite 3-methoxytyramine (3-MT) may be a useful index of DA release in vivo. In the present study, in vivo microdialysis was utilized to assess directly the relationship between extracellular DA and 3-MT in the striatum of rats following a variety of pharmacological manipulations. Apomorphine, a DA receptor agonist, produced a rapid, transient decrease in both DA and 3-MT. Conversely, the DA receptor antagonist haloperidol produced a concomitant increase in extracellular DA and 3-MT. Increases in DA and 3-MT were also noted following the administration of the DA uptake inhibitor, bupropion. Local application of tetrodotoxin resulted in the complete elimination of measurable amounts of DA and 3-MT in the dialysate, gamma-Butyrolactone also greatly decreased DA and 3-MT. Finally, d-amphetamine produced a large increase in DA and 3-MT in animals that had been treated previously with gamma-butyrolactone. The Pearson correlation coefficients for DA and 3-MT following these manipulations ranged from 0.87 to 0.97. These data indicate that interstitial 3-MT is an accurate index of DA release. However, when compared with previous ex vivo findings, the present results also suggest that changes in tissue concentrations of 3-MT may not reliably reflect DA release following certain pharmacological manipulations.     

3-Methoxytyramine Is the Major Metabolite of Released Dopamine in the Rat Frontal Cortex: Reassessment of the Effects of Antipsychotics on the Dynamics of Dopamine Release and Metabolism in the Frontal Cortex, Nucleus Accumbens, and Striatum by a Simple Two Pool Model

Abstract: 3-Methoxytyramine (3-MT) and 3,4-dihydroxyphenylacetic acid (DOPAC) rates of formation were used, respectively, to assess the dynamics of dopamine (DA) release and turnover in the rat frontal cortex, nucleus accumbens, and striatum. Assuming total (re)uptake and metabolism of released DA are relatively uniform among the three brain regions, a simplified two pool model was used to assess the metabolic fate of released DA. Under basal conditions, 3-MT formation was found to comprise >60% of total DA turnover (sum of 3-MT plus DOPAC rates of formation) in the frontal cortex, and not more than 15% in the nucleus accumbens and striatum. Haloperidol increased the 3-MT rate of formation to a greater extent in the frontal cortex than in the two other regions. Clozapine increased the 3-MT rate of formation in the frontal cortex and decreased it in the striatum. Both drugs increased DOPAC rate of formation in the frontal cortex and nucleus accumbens. It was elevated by haloperidol but not clozapine in the striatum. It is concluded that (1) O -methylation is a prominent step in the catabolism of DA in the frontal cortex under both physiological conditions and after acute treatment with antipsychotics, (2) 3-MT is the major metabolite of released DA in the frontal cortex and possibly also in the nucleus accumbens and striatum, (3) in contrast to the frontal cortex, most of the DOPAC in the nucleus accumbens and striatum appear to originate from intraneuronal deamination of DA that has not been released, (4) because presynaptic uptake and metabolism of DA give rise to DOPAC, whereas postsynaptic uptake and metabolism produced both DOPAC and 3-MT, the ratio of 3-MT to DOPAC rates of formation can be a useful index of reuptake inhibition.     

Free and conjugated catecholamines and metabolites in cat urine after hypoxia

Catecholamine and metabolite excretion was studied in the cat after 6 h of 7.5% O2 hypoxia. Norepinephrine (NE) release from sympathetic nervous endings was strongly activated, whereas epinephrine (E) excretion was only slightly increased. A noteworthy result was the increase of dopamine (DA) and its metabolites [3-methoxytyramine (MT); 3,4-dihydroxyphenylacetic acid (DOPAC)] in urine samples. This increased release does not seem to originate from the central nervous system, but rather from peripheral dopaminergic structures; available knowledge on peripheral DA suggests that the hypoxia-induced DA release might be partly related to chemosensory or renal function. Indeed, in addition to enhanced DA and NE excretion, we observed an increase in sodium excretion that correlated with both DA and NE. Analysis of free and conjugated urinary metabolites showed that only free NE and both free and conjugated normetanephrine were increased in urine after hypoxic stress. Among DA metabolites, conjugated DOPAC was the main DA metabolite in the basal state and after hypoxia. Both the free and the conjugated forms of DA, MT, and DOPAC were increased by hypoxia.     

On the Significance of Endogenous 3-Methoxytyramine for the Effects of Centrally Acting Drugs on Dopamine Release in the Rat Brain

Abstract: 3-Methoxytyramine (3-MT) was measured in the striata of rats killed by microwave radiation. Apomorphine, γ-butyrolactone (GBL), and reserpine decreased the 3-MT content. A slight but transient increase in 3-MT was observed after haloperidol. The turnover rate of 3-MT was unchanged 60 min after haloperidol treatment. (+)-Amphetamine induced a pronounced rise in the 3-MT content, which was potentiated after combined treatment with haloperidol. The increased 3-MT turnover rate that was observed after amphetamine treatment suggests that monoamine oxidase (MAO) inhibition is no explanation for the mechanism of interaction of this drug with dopamine (DA) metabolism. The central stimulants amphonelic acid and nomifensine in-creased 3-MT levels; no substantial change was seen after benztropine, morphine, or oxotremorine. It is concluded that a decreased release of DA is closely and rapidly reflected by decreased formation of 3-MT. 3-MT seems to be a much better indicator for decreased DA release than 3,4-dihydroxyphenylacetic acid or homovanillic acid.     

Effect of submaximal muscular exercise of short duration on urinary excretion of catecholamines, DOPA and their metabolites]

Thirteen human subjects were submitted to a moderate muscular work on ergometric bicycle (at intensity corresponding to 80% of maximal oxygen uptake during 10 min). No modifications were observed in the urinary amounts of the three catecholamines (A, NA, DA), DOPA, DOPAC and 3-MT. On the contrary, the excretion of metadrenaline (MN) and normetadrenaline (NMN) was slightly increased, showing a mild stimulation of adrenergic system. Our result point out the interest of urinary methoxyamines as useful index of adrenergic activity in man. For experimental and physiopathological use, the metabolic alteration induced by a short submaximal muscular work is negligible for most adrenergic compounds, except for MN and NMN, the amounts of which are slightly modified.     

The Dopamine Metabolite 3-Methoxytyramine Is Not a Suitable Indicator of Dopamine Release in the Rat Brain

It has been postulated that changes in the concentration of 3-methoxytyramine (3-MT) in the brain might reflect changes in the release of 3,4-dihydroxyphenylethylamine (DA, dopamine) and, therefore, might be used as an index of dopaminergic activity in the brain. 3-MT is known to accumulate rapidly after death. Killing by microwave irradiation (MWR) is considered to be the method of choice to obtain "undisturbed" 3-MT concentrations. We measured striatal 3-MT concentrations even lower than those following MWR when the brains were excised and frozen in dry ice very rapidly (typical time between decapitation and freezing of the brain 22 s). There was a linear increase in striatal 3-MT concentration when the time between decapitation and freezing was varied between 13 and 300 s. Extrapolation to time zero indicated negligible amounts of 3-MT at the time of decapitation. In addition, it was observed that DA, 3,4-dihydroxyphenylacetic acid, and homovanillic acid decompose during the cooling phase after heating the brain by microwave. It is concluded that MWR induces artifactual changes in the postmortem levels of DA and metabolites. Consequently 3-MT cannot be considered to be a reliable indicator of DA release in the rat brain.     

Intracerebral dialysis: direct evidence for the utility of 3-MT measurements as an index of dopamine release

Intracerebral dialysis was used to monitor the in vivo efflux of striatal dopamine (DA), homovanillic acid (HVA), dihydroxyphenylacetic acid (DOPAC) and 3-methoxytyramine (3-MT) in the pentobarbital anesthetized rat. In untreated rats, there were low levels of extra-cellular DA and 3-MT which were increased 15-fold by treatment with amphetamine. Under basal and drug-stimulated conditions, 3-MT concentrations were maintained at approximately 30% of the extracellular DA levels. These data agree with in vivo turnover estimates which indicate that 20 to 30% of DA turnover is through the 3-MT pool in the striatum. In contrast, extracellular DOPAC and HVA levels were reduced only slightly by amphetamine and with a delayed onset. Our data support the hypothesis that striatal DOPAC is an accurate index of intraneuronal DA metabolism and that 3-MT is an index of the extracellular concentration of DA.     

Role of the nitric oxide/cyclic GMP pathway and extracellular environment in the nitric oxide donor-induced increase in dopamine secretion from PC12 cells: a microdialysis in vitro study

In vitro microdialysis was used to investigate the mechanism of nitric oxide (NO) donor-induced changes in dopamine (DA) secretion from PC12 cells. Infusion of the NO-donor S-nitroso-N-acetylpenicillamine (SNAP, 1.0 mm) induced a long-lasting increase in DA and 3-methoxytyramine (3-MT) dialysate concentrations. SNAP-induced increases were inhibited either by pre-infusion of the soluble guanylate cyclase (sGC) inhibitor 1H-[1,2,4] oxadiazolo[4,3]quinoxalin-1-one (ODQ, 0.1 mm) or by Ca2+ omission. Ca2+ re-introduction restored SNAP effects. SNAP-induced increases in DA + 3-MT were unaffected by co-infusion of the l-type Ca2+ channel inhibitor nifedipine. The NO-donor (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide (NOR-3, 1.0 mm) induced a short-lasting decrease in dialysate DA + 3-MT. Ascorbic acid (0.2 mm) co-infusion allowed NOR-3 to increase dialysate DA + 3-MT. ODQ pre-infusion inhibited NOR-3 + ascorbic acid-induced DA + 3-MT increases. Infusion of high K+ (75 mm) induced a 2.5-fold increase in dialysate DA + 3-MT. The increase was abolished by NOR-3 co-infusion. Conversely, co-infusion of ascorbic acid (0.2 mm) with NOR-3 + high K+ restored high K+ effects. Co-infusion of nifedipine inhibited high K+-induced DA + 3-MT increases. These results suggest that activation of the NO/sGC/cyclic GMP pathway may be the underlying mechanism of extracellular Ca2+-dependent effects of exogenous NO on DA secretion from PC12 cells. Extracellular Ca2+ entry may occur through nifedipine-insensitive channels. NO effects and DA concentrations in dialysates largely depend on both the timing of NO generation and the extracellular environment in which NO is generated.     

Relationship Between Catechol-O-Methyltransferase and Phenolsulfotransferase in the Metabolism of Dopamine in the Rat Brain

The relationship between phenolsulfotransferase (PST) and catechol-O-methyltransferase (COMT) in the metabolism of free 3,4-dihydroxyphenylethylamine (DA, dopamine) in the rat brain was studied. In rats not pretreated with a monoamine oxidase (MAO) inhibitor a huge increase of free DA in the brain, following an intraperitoneal injection of L-3,4-dihydroxyphenylalanine (L-DOPA) or an intraventricular injection of free DA, did not lead to any noticeable change in DA sulfate or 3-methoxytyramine (3-MT), which remained undetectable by the present HPLC method. However, in rats previously treated with the MAO inhibitors pargyline or tranylcypromine, the same L-DOPA or free DA treatment resulted in significant increases in both 3-MT and DA sulfate in the hypothalamus, brainstem, and striatum. This response of COMT and PST was not affected by prior treatment of the rats with 6-hydroxydopamine, which suggests that O-methylation and sulfoconjugation occur outside adrenergic neurons not destroyed by the neurotoxin. Inhibition of COMT activity did not lead to any increase in DA sulfate, which showed that despite their common mode of action (both enzymes react preferentially at the same hydroxyl group in the DA molecule), the two enzymes are not competitive. After MAO inhibition there were strong correlations between an increase in DA sulfate and 3-MT on the one hand, and between free DA and 3-MT on the other. Because 3-MT is a marker of central DA release, these data suggest that inhibition of MAO activity not only affects DA metabolism by this enzyme but also influences DA release in the rat brain.     

Catecholamine Glucuronidation: An Important Metabolic Pathway for Dopamine in the Rat

Abstract: In the present study, we found that large quantities of dopamine (DA) glucuronide were present in rat cerebrospinal fluid (CSF), plasma, and urine, whereas the glucuronides of norepinephrine (NE) and epinephrine (E) were almost undetectable. The high urinary excretion of DA glucuronide was in a range comparable to that of homovanillic acid (HVA). Sulfates of DA, NE, and E were measurable in all three body fluids, but only in small quantities. The measured DA glucuronide was predominantly of endogenous origin, as the feeding of sucrose instead of routine diet did not reduce the urinary output of DA glucuronide. Adrenalectomy but not peripheral sympathectomy induced by chronic guanethidine injection substantially decreased plasma DA glucuronide concentrations, indicating that the adrenals serve as an important source of endogenous DA glucuronide. The data suggest that glucuronidation constitutes an important metabolic pathway for endogenous DA of central and peripheral origin in rats; this route, however, is exclusive to DA and appears to play a negligible role for NE and E.     

Effects of dopamine metabolites on locomotor activities and on the binding of dopamine: relevance to the side effects of L-dopa

L-dopa is the major treatment for Parkinson's disease (PD), but its efficacy is limited by the presence of dyskinesia. The dyskinesia develops over a period of exposure to L-dopa and is related to the dosage, therefore, the cause may involve inductive changes that produce toxic levels of metabolites, interfering with dopamine (DA) neurotransmission. Chronic L-dopa induces catechol-O-methyltransferase (COMT) and methionine adenosyl transferase (MAT), enzymes involved in the methylation of catecholamines (CA). In addition, high levels of 3-O-methyl-dopa have been reported in the plasma of dyskinetic PD patients, treated with L-dopa, as compared to non-dyskinetic patients, therefore, the methyl metabolites of CA may be increased during L-dopa therapy and may be involved in the dyskinesia. Since large amounts of DA are produced from L-dopa, and DA is extensively methylated, the methyl metabolites of DA, 3-methoxytyramine (3-MT) and 3,4-dimethoxyphenylethylamine (DIMPEA), may be also involved. The first step in knowing this, is to assess the behavioral and DA-receptor activities of 3-MT and DIMPEA. In the rat, the intraventricular injection of 0.5 micromol of DIMPEA increased the total distance traveled (TD) by over 100%, the number of movement (NM) made by 40% and the time spent moving (MT) by about 36%. Identical doses of 3-MT decreased the TD by 42%, NM by 22% and MT by 39%. DIMPEA (1 mM) increased the binding of DA with brain membranes by 44.7%, whereas 3-MT decreased it by 15.8%. The results show that 3-MT and DIMPEA are behaviorally active, and in parallel, they interact with the binding sites for DA, consequently, they may contribute to the side effects of L-dopa. L-dopa produces high levels of DA and induces MAT and COMT. It is proposed, therefore, that DA will be methylated to 3-MT and 3-MT to DIMPEA. At threshold level each product will inhibit, allosterically, its enzyme of methylation, causing sequential and rhythmic up and down regulation of its concentration. At peak levels these hydrophobic metabolites will modulate the actions of DA on synaptic membranes, causing abnormal movements, at times, resembling the "on-off effects".     

Suppression of norepinephrine secretion by dopamine in children with neuroblastoma: evidence for precursor inhibition in catecholamine pathway

To elucidate catecholamine (CA) secretory dynamics in neuroblastoma, urinary excretion of CAs and their metabolites was serially measured in 6 patients aged 3 months to 3 years before and during treatment. After tumor extirpation, increased urinary CAs were promptly normalized; the reduction reflected the amount of CA production from the tumor. Urinary dopamine (DA) showed the most prominent reduction, whereas DA content in the tumor was very small, indicating that the DA produced was immediately released from the tumor and metabolized in extra-tumor tissues. In contrast, patients receiving chemotherapy continued to excrete excess DA and homovanillic acid (HVA), which were increased further at recidivation. One patient showed an inverse correlation between DA and norepinephrine (NE) excretion; a decrease in DA was associated with an increase in NE and plasma DA-beta-hydroxylase (DBH) activity. A similar inverse correlation was also noted between NE and vanillylmandelic acid (VMA) or 3-methoxy-4-hydroxyphenylglycol (MHPG) excretion, while HVA and dihydroxyphenylacetic acid (DOPAC) were positively correlated with DA excretion. Urinary HVA and VMA were lineally correlated but in a patient excreting an enormous amount of DA, urinary VMA was markedly suppressed in terms of HVA excretion. Excessive DA induced an increase in renal water output but did not enhance Na and K excretion. These results indicate that endogenous DA overload in neuroblastoma inhibits NE production by suppressing DBH activity as well as by forming VMA and MHPG. This precursor regulation appears to be the characteristic of the CA metabolic pathway.     

Plasma catecholamines and urinary excretion of their main metabolites in three models of portal hypertension

We have measured, by a specific radioenzymoassay, the plasma concentration of dopamine (DA) and norepinephrine (NE) and by gas chromatography the urinary excretion of some catecholamine metabolites (HVA, homovanillic acid, DOPAC, dihydroxyphenyl acetic acid; VMA, vanilmandelic acid, and DOPEG, dihydroxyphenyl glycol) in three groups of rats with portal hypertension: cirrhotic rats (CR), rats with progressive portal hypertension (PPH) and rats with progressive hepatic congestion (PHC). The three groups of rats had portal hypertension. PPH and PHC had also intrahepatic hypertension. CR rats showed an increased urinary excretion of NE and DA metabolites with a normal plasma concentration of these catecholamines, suggesting an increased turnover of NE and DA in this experimental model. PPH animals had a high plasma DA concentration with a decreased urinary excretion of catecholamine metabolites. PHC showed high plasma DA and NE levels with normal or increased urinary excretion of its metabolites. These results suggest that an increased neural activity is present in the early stages of experimental cirrhosis in rats and this alteration does not seem directly related to the portal hypertension but perhaps to the intrahepatic hypertension or to the hepatocellular damage.     

Turnover of Dopamine and Dopamine Metabolites in Rat Brain: Comparison Between Striatum and Substantia Nigra

Measurements of the turnover of dopamine (DA) and DA metabolites have been performed in the striatum and substantia nigra (SN) of the rat. Turnover rates of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid have been assessed from the disappearance rates after blocking their formation by inhibition of monoamine oxidase by pargyline and of catechol-O-methyltransferase by tropolone. DA turnover has been measured as 3-methoxytyramine (3-MT) plus DA accumulation rate after MAO inhibition by pargyline and as accumulation rate of 3,4-dihydroxyphenylalanine (DOPA) after inhibition of aromatic amino acid decarboxylase by NSD 1015 or NSD 1034. These measures of DA turnover have been compared with alpha-methyl-p-tyrosine (alpha-MT)-induced DA disappearance rate. In SN all the different measures of DA turnover are in the same range (55-62 nmol/g protein/h) whereas in striatum DOPA accumulation rate after NSD 1015 and alpha-MT-induced DA disappearance rate (16-23 nmol/g/h) are much lower than DOPAC disappearance rate after pargyline, 3-MT plus DA accumulation rate after pargyline, and DOPA accumulation rate after NSD 1034 (39-46 nmol/g/h). The data confirm our previous findings indicating that the fractional turnover rate of DA is more rapid in SN than in striatum and that O-methylation of DA is relatively more important in SN. In striatum at least two pools of DA with different turnover rates appear to exist, whereas in SN, DA behaves as if located in a single compartment.     

Relationship Between Plasma and Cerebrospinal Fluid Norepinephrine and Dopamine Metabolites in a Nonhuman Primate

Major and minor pathways of metabolism in the mammalian CNS result in the formation of 3-methoxy-4-hydroxyphenylethylene glycol (MHPG) and normetanephrine (NMN) from norepinephrine (NE), and homovanillic acid (HVA) and 3-methoxytyramine (3-MT) from dopamine (DA), respectively. The correlational relationships between HVA and 3-MT and between MHPG and NMN in primate CSF and plasma have not been described. These relationships may help to elucidate the usefulness of CSF and plasma metabolites as indices of CNS NE and DA activity. In addition, because NMN is unlikely to cross the blood-brain barrier. CSF NMN concentrations would not be confounded by contributions from plasma, which is a major issue with CSF MHPG. We have obtained repeated samples of plasma and CSF from drug-naive male squirrel monkeys and have measured the concentrations of MHPG, HVA, NMN, and 3-MT to define their correlational relationships. For the NE metabolites, significant correlations were obtained for CSF MHPG and NMN (r = 0.806, p less than 0.001), plasma MHPG and CSF NMN (r = 0.753, p less than 0.001), and plasma and CSF MHPG (r = 0.776, p less than 0.001). These results suggest that CSF and plasma MHPG and CSF NMN may reflect gross changes in whole brain steady-state noradrenergic metabolism. Only a single significant relationship was demonstrated for the DA metabolites, with CSF 3-MT correlating with plasma HVA (r = 0.301, p less than 0.025). The results for the DA metabolites probably reflect regional differences in steady-state brain dopaminergic metabolism.     

In vivo release of endogenous dopamine, 5-hydroxytryptamine and some of their metabolites from rat caudate nucleus by phenylethylamine

The in vivo release of endogenous 3,4-dihydroxyphenylethylamine (DA) and its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 3-methoxytyramine (3-MT), and of 5-hydroxytryptamine (5-HT) and its metabolite, 5-hydroxyindoleacetic acid (5-HIAA), has been measured in the caudate nucleus of the anesthetized rat. A push-pull cannula was implanted into the brain, and the tissue perfused with artificial CSF or artificial CSF containing 5×10^–4 M phenylethylamine. The perfusate was collected and analyzed for DA, 5-HT and their metabolites by high performance liquid chromatography with electrochemical detection (HPLC-ECD). DA was released by phenylethylamine at rates significantly greater than its basal rate. 3-MT and 5-HT were undetectable in perfusates collected under basal conditions, but could be detected readlly during phenylethylamine stimulation. DOPAC, HVA and 5-HIAA concentrations were not significantly affected by phenylethylamine. The results suggest (1) that phenylethylamine may exert its behavioural effects through increased release of both DA and 5-HT, and (2) that in vivo measurements of the acid metabolites alone may not be indicative of the release of the amines.Special Issue Dedicated to Dr. Abel Lajtha.     

Inhibition of brain mitochondrial respiration by dopamine and its metabolites: implications for Parkinson's disease and catecholamine-associated diseases

A structure-potency study examining the ability of dopamine (DA), its major metabolites and related amine and acetate congeners to inhibit NADH-linked mitochondrial O(2) consumption was carried out to elucidate mechanisms by which DA could induce mitochondrial dysfunction. In the amine studies, DA was the most potent inhibitor of respiration (IC(50) 7.0 mm) compared with 3-methoxytryramine (3-MT, IC(50) 19.6 mm), 3,4-dimethoxyphenylethylamine (IC(50) 28.6 mm), tyramine (IC(50) 40.3 mm) and phenylethylamine (IC(50) 58.7 mm). Addition of monoamine oxidase (MAO) inhibitors afforded nearly complete protection against inhibition by phenylethylamine, tyramine and 3,4-dimethoxyphenylethylamine, indicating that inhibition arose from MAO-mediated pathways. In contrast, the inhibitory effects of DA and 3-MT were only partially prevented by MAO blockade, suggesting that inhibition might also arise from two-electron catechol oxidation and quinone formation by DA and one-electron oxidation of the 4-hydroxyphenyl group of 3-MT. In the phenylacetate studies, 3,4-dihydroxyphenylacetic acid (DOPAC) was equipotent with DA in inhibiting respiration (IC(50) 7.4 mm), further implicating the catechol reaction as the cause of inhibition. All other carboxylate congeners; phenylacetic acid (IC(50) 13.0 mm), 4-hydroxyphenylacetic acid (IC(50) 12.1 mm), 4-hydroxy-3-methoxyphenylacetic acid (HVA, IC(50) 12.0 mm) and 3,4-dimethoxyphenylacetic acid (IC(50) 10.2 mm), were equipotent respiratory inhibitors and two- to fourfold more potent than their corresponding amine. These latter findings suggest that the phenylacetate ion can also contribute independently to mitochondrial inhibition. In summary, mitochondrial respiration can be inhibited by DA and its metabolites by four distinct MAO-dependent and independent mechanisms.     

Dopamine metabolism and free-radical related mitochondrial injury during transient brain ischemia in gerbils

Regional extracellular release of dopamine (DA) and its metabolites, 3,4-dihydroxy-phenylacetic acid (DOPAC), homovanillic acid (HVA) and 3-methoxytyramine (3-MT) was measured in gerbils (with or without pargyline pretreatment) subjected to bilateral carotid artery occlusion (15 min) and various periods of recirculation (up to 6 hr), utilizing intracerebral microdialysis and high-performance liquid chromatography (HPLC) with electrochemical detection. Mitochondrial monoamine oxidase (MAO) and superoxide dismutase (SOD) activities andin vitro stimulated lipid peroxidation (TBARM) were determined in separate experimental groups of animals. The ischemically induced DA release, decrease of MAO-derived DA metabolites DOPAC and HVA, and accumulation of 3-MT were potentiated and prolonged by pargyline pretreatment. Mitochondrial MAO and SOD activities were significantly reduced during ischemia alone and up to 1 hr of reperfusion, whereas TBARM was enhanced during reflow only. The data suggest that reduced activity of mitochondrial antioxidative enzyme(s) but not DA metabolism by MAO may contribute to free radical-mediated injury of (mitochondrial) membranes.     

Urinary excretion of bioactive amines and their metabolites in psychiatric patients receiving phenelzine

Phenelzine [2-phenylethylhydrazine] (PLZ), a potent inhibitor of monoamine oxidase (MAO)-A and-B, is used widely in psychiatry. We have studied the effects of PLZ administration on urinary excretion of several bioactive amines and their metabolites in psychiatric patients. Urine samples (24-hour) were collected prior to treatment and again at 2 and 4 weeks of treatment with PLZ (30–90 mg daily in divided doses). Amines and metabolites analyzed included 2-phenylethylamine (PEA), m-and p-tyramine (m-and p-TA), phenylacetic acid (PAA), m-and p-hydroxyphenylacetic acid (m-and p-OH-PAA), tryptamine (T), 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), normetanephrine (NME), 3-methoxy-4-hydroxyphenylglycol (MHPG), 3-methoxytyramine (3-MT), and homovanillic acid (HVA). Levels of PEA, p-TA, 5-HT, and T were elevated during treatment with PLZ, but no significant changes in urinary excretion of the acid metabolites PAA, p-OH-PAA, and 5-HIAA were observed. Urinary levels of the noradrenaline metabolites NME and MHPG were increased and decreased, respectively; a similar pattern was observed with the dopamine metabolites 3-MT and HVA. There was an elevation in levels of m-TA and a decrease in its acid metabolite m-OH-PAA during the treatment with PLZ.     

In Vivo Assessment of Dopamine and Norepinephrine Release in Rat Neocortex: Gas Chromatography-Mass Spectrometry Measurement of 3-Methoxytyramine and Normetanephrine

A new method with the sensitivity and specificity required to measure regional levels of 3-methoxytyramine (3-MT) and normetanephrine (NMN) in the rat cortex is described. The method utilizes a liquid ion exchanger to isolate the parent amines, dopamine (DA) and norepinephrine (NE), along with their methylated metabolites. These samples are derivatized and analyzed by negative ion gas chromatography-mass spectrometry. Using this method, we examined a number of drug actions on steady-state levels as well as pargyline-induced increases in 3-MT and NMN. In the prefrontal cortex, cingulate cortex, striatum, and olfactory tubercle, nomifensine was found to increase 3-MT steady-state levels and accumulation rates. Similar actions of this drug were observed in the cingulate and prefrontal cortices with NMN. In contrast, clonidine decreased cortical NMN levels and accumulation. A unique action was observed with haloperidol, in that both 3-MT levels and accumulation after pargyline were increased in the nigrostriatal and mesolimbic dopaminergic projections, whereas only the accumulation rates were accelerated in the mesocortical projections. In summary, our data indicate that this new assay is a useful approach for the in vivo evaluation of DA and NE release in cortical regions of the rat. This approach is unique in that no surgery, restraint, or anesthetic is required, thereby permitting more complicated experimental paradigms to be utilized.