Amino Acid Decarboxylase Inhibitor Research Articles

GM1 ganglioside rescues substantia nigra pars compacta neurons and increases dopamine synthesis in residual nigrostriatal dopaminergic neurons in MPTP-treated mice.

GM1 ganglioside has been shown to stimulate recovery of the damaged dopamine system under a number of different circumstances. In addition to rescue of damaged dopamine neurons, the present study assessed the ability of GM1 to enhance the synthesis of dopamine in remaining nigrostriatal neurons following 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) exposure. There was a significantly greater accumulation of L-dopa 30 min after aromatic amino acid decarboxylase inhibition with NSD-1015 (100 mg/kg) and an increase in the ratio of L-dopa to dopamine in MPTP+GM1-treated mice than in mice that received only MPTP. This effect of GM1 on dopamine synthesis was dependent upon the degree of initial damage to the nigrostriatal dopamine system. That is, the GM1 effect on dopamine synthesis could not be demonstrated in mice with greater than 95% striatal dopamine loss and 75% substantia nigra dopamine neuron loss. These results suggest that in addition to previously reported effects of GM1 on rescue and repair of dopaminergic neurons, GM1 may also have the ability to enhance dopamine synthesis in residual dopaminergic neurons. Direct effects on dopamine neurochemistry may contribute to functional improvement seen after GM1 treatment in various models of parkinsonism.

Infertility in Transgenic Mice Overexpressing the Bovine Growth Hormone Gene: Disruption of the Neuroendocrine Control of Prolactin Secretion during Pregnancy1

Transgenic female mice overexpressing the bovine growth hormone (bGH) gene with the phosphoenolpyruvate carboxykinase (PEPCK) promoter exhibit severe reproductive deficits. Although these animals ovulate and conceive normally, pregnancy is arrested due to luteal failure, leading to the loss of embryos during early gestation. The results of replacement therapy suggested that luteal failure was secondary to prolactin (PRL) deficiency. The objective of this study was to examine the neuroendocrine control of PRL secretion during early pregnancy in PEPCKbGH-1 transgenic mice. Normal and transgenic littermates were killed by decapitation on Day 7 postcoitum (p.c.) at 1500, 1800, or 2100 h, i.e., the period including the expected diurnal PRL surge in pregnant mice. In normal females, plasma PRL levels were significantly elevated at 1800 h when compared to the levels measured at 1500 or 2100 h, but no temporal variation in PRL levels was found in transgenic mice. In normal females, the content of dopamine in the median eminence was reduced at 1800 h, i.e., at the time of the PRL surge. In contrast, no temporal changes were detected in the median eminence dopamine content in transgenic mice. Twice-daily injections of domperidone, a dopamine receptor blocker, increased the incidence of pregnancy in transgenic females. After treatment with aromatic amino acid decarboxylase inhibitor NSD-1015 on Day 7 p.c., plasma levels of PRL were similarly elevated in transgenic and normal females. However, the accumulation of 5-hydroxytryptophan (5-HTP) in the medial basal hypothalamus after this treatment was significantly smaller in transgenic than in normal females.(ABSTRACT TRUNCATED AT 250 WORDS)

Medical treatment of Parkinson's disease.

Parkinson’s disease (PD) is a progressive neurodegenerative disease. It’s main feature is the nigrostriatal neuronal cell loss resulting in striatal dopamine (DA) deficiency. L-dopa + peripheral amino acid decarboxylase inhibitor (DDI: Sinemet or Madopar) is still the most effective agent for the symptomatic treatment of PD, but the dopa therapy is far from perfect. For advanced cases especially for patients with wearing off of the drug effect, fluctuation of response and “on-off, ” the conventional preparation such as Sinemet or Madopar due to their short T1/2 is no longer suitable for clinical use. Thus controlled release preparation e.g. Sinemet CR or Madopar HBS is preferable.KeywordsStriatal DopamineAdvanced CaseCOMT InhibitorProgressive Neurodegenerative DiseasePsychiatric ComplicationThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Regulation by D2 Dopamine Receptors of in Vivo Dopamine Synthesis in Striata of Rats and Mice with Experimental Parkinsonism

The effects of D2 dopamine (DA) receptor antagonism or stimulation by systemic haloperidol or quinpirole, respectively, on in vivo DA synthesis in 6-hydroxydopamine (6-OHDA)-lesioned rats and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)treated black mice were assessed by measuring the rate of dihydroxyphenylalanine (DOPA) accumulation following acute inhibition of l-aromatic amino acid decarboxylase with NSD-1015. 6-OHDA and MPTP caused partial lesions of nigrostriatal input to the striatum. Dopamine synthetic capacity was preserved relative to the severity of nigrostriatal lesion over a broad range of DA depletions. An exponential increase in fractional DA synthesis (the ratio DOPA/DA) was observed with increasing DA depletion, suggesting an elevation of the DA synthetic capacity per surviving DA terminal. In both lesioned rats and mice, haloperidol caused a significant increase in fractional DA synthesis above that induced by the lesion alone, while quinpirole significantly depressed fractional DA synthesis. Our results provide evidence that nigrostriatal terminals acquire increased DA synthetic capacity as nigrostriatal lesions exceed 90%, but that the increase in fractional DA synthesis observed in partially lesioned animals is not due to a loss of autoreceptor function. Pharmacological strategies to stimulate DA synthesis and release in moderately advanced Parkinson's disease should be pursued.

Enzymes for the resolution of α-tertiary-substituted carboxylic acid esters

Aromatic α-amino-α-methyl acids and α-hydrazino-α-methyl acids are known aromatic amino acid decarboxylase inhibitors. Specific derivatives such as 2-amino-2-methyl-3-(3,4-dihydroxyphenyl)propanoate, Aldomet®, and 2-hydrazino-2-methyl-3-(3,4-dihydroxypheny Lodosyn®, have been developed as therapeutic agents to treat hypertension and Parkinson's disease, respectively. We recently reported a method for the kinetic resolution of the racemic esters of such compounds using a crude preparation of a novel enzyme catalyst from the yeast Candida lipolytica (Yee, C.; Blythe, T. A.; McNabb, T. J.; Walts, A. E. J. Org. Chem. 1992, 57, 3525-3527). Here we report the purification and initial characterization of the active enzyme component, an enzyme given the name Candida lipolytica ester hydrolase (CLEH). CLEH was purified to > 95% homogeneity by chromatography on Matrex TM Blue B resin. The enzyme was found to be a glycoprotcin with M r = 80,000–300,000. In addition to esterolytic activity, the enzyme was found to catalyze the hydrolysis of amides, anilides and peptides. Sequence analysis of internal peptides of CLEH revealed striking homology to a number of enzymes belonging to the group of serine carboxypeptidases (E.G. 3.4.16.1). One peptide aligned with the canonical serine carboxypeptidase active site sequence, GESYAG. Based on the structural relationship of CLEH to serine carboxypeptidases, three representative serine carboxypeptidases were evaluated for their utility in resolving racemic α-tertiary ester substrates and compared with the activity of CLEH. All enzymes revealed similarly high activity and enantioselectivity towards the α-hydrazino-α-methyl ester precursor of the Parkinson-drug Carbidopa. However, differences in enantioselectivity were observed with other α-tertiary-substituted ester substrates. Serine carboxypeptidase-catalyzed ester resolutions thus offer a new route to many sterically hindered homochiral α-amino, α-hydrazino and α-hydroxy carboxylic acids.

The effect of short-photoperiod exposure on tuberoinfundibular dopamine neurons in male and female Syrian hamsters.

In male hamsters, exposure to a short photoperiod results in a significant decrease in median eminence (ME) dopamine (DA) concentrations. The mechanism responsible for this decrease in DA is unknown. The experiments described in this paper were designed to examine the effects of photoperiod on DA metabolism and synthesis in the ME to determine if a change in these processes is responsible for the short-photoperiod-induced decrease in ME DA concentrations. In the first experiment, the metabolism of DA in tuberoinfundibular dopamine (TIDA) neuronal terminals was determined by measuring ME concentrations of 3,4-dihydroxyphenylacetic acid (DOPAC; a major metabolite of DA) and DA in male and female hamsters housed in long and short photoperiods. In both males and females, exposure to the short photoperiod induced a collapse of the reproductive system and a reduction in circulating prolactin. In males, but not in females, exposure to the short photoperiod reduced ME DA concentrations; however, DOPAC concentrations were not affected by photoperiod. Thus, the decrease in ME DA seen in males is not the result of an increase in DA metabolism. In the second experiment, tyrosine hydroxylase (TH) activity in the ME of males was determined by injecting animals housed in long and short photoperiods with a L-aromatic amino acid decarboxylase inhibitor (NSD 1015) and measuring 3,4-dihydroxyphenylalanine (DOPA). Consistent with Experiment 1, ME DA concentrations were significantly decreased in gonadally regressed males housed in a short photoperiod; however, ME DOPA accumulation was not affected. Thus, the observed decrease in DA is not the result of a decrease in TH activity in the ME. The results of the experiments presented here indicate that (1) in males but not females, the decrease in circulating prolactin seen in animals housed in a short photoperiod for 12 weeks is associated with a decrease in ME DA concentrations, and (2) the decrease in ME DA seen in males housed in a short photoperiod is not the result of an increase in DA metabolism or a decrease in synthesis by TIDA neurons.

Evaluation of 3-[ 18F]Fluoro-α-fluoromethyl- p-tyrosine as a tracer for striatal tyrosine hydroxylase activity

3-[ 18F]Fluoro-α-fluoromethyl- p-tyrosine (3-F-FMPT) was evaluated as a tracer for CNS tyrosine hydroxylase (TH) activity in rodents and in a rhesus monkey. Results of in vitro experiments using rat striatal homogenates showed that the introduction of fluorine into the 3-phenyl position did not significantly alter the ability of FMPT to act as a TH-activated l-aromatic amino acid decarboxylase ( l-AAAD) inhibitor. These studies further showed that 3-F-FMPT-induced l-AAAD inhibition was dose-dependent. Furthermore, striatal homogenates prepared from rats pretreated with the potent TH inhibitor α-methyl- p-tyrosine was found to have diminished 3-F-FMPT-induced l-AAAD inhibition. However, despite these promising in vitro results, the biodistribution of this compound in mice showed low brain uptake and fast clearance through the kidneys. A PET study using a Rhesus monkey injected with 3-[ 18F]F-FMPT confirmed the results obtained in mice, i.e. negligible brain uptake but high localization in the bladder. We conclude that 3-[ 18F]F-FMPT would not be useful as a tracer for cerebral TH activity.

The noradrenaline precursor l‐threo‐3,4‐dihydroxyphenylserine exhibits antinociceptive activity via central α‐adrenoceptors in the mouse

1. Systemic (s.c. or p.o.) administration of L-threo-3,4-dihydroxyphenylserine (droxidopa, L-threo-DOPS; L-DOPS), a noradrenaline precursor, at a dose-range of 100-800 mg kg-1, produced naloxone-resistant antinociception in a dose-dependent manner in the mouse, as assessed by the tail flick test, kaolin-induced writhing test and formalin-induced nociception test. 2. Antinociception elicited by L-DOPS (400 mg kg-1, s.c.) was not affected by s.c. injection of benserazide, a peripherally preferential L-aromatic amino acid decarboxylase inhibitor, but was suppressed by its intracerebroventricular (i.c.v.) injection. 3. I.c.v. or intrathecal (i.t.) administration of the non-selective alpha-blocker, phentolamine, significantly reduced L-DOPS-induced antinociception. 4. I.c.v. administration of the alpha 1-blocker, prazosin, but not the alpha 2-blocker, yohimbine, abolished the antinociceptive effects of L-DOPS. In contrast, both blockers, when administered i.t., exhibited significant inhibitory effects. 5. These results suggest that systemic L-DOPS produces opioid-independent antinociception, mediated by supraspinal alpha 1-adrenoceptors and by spinal alpha 1- and alpha 2-adrenoceptors and may predict additional therapeutic applications of L-DOPS as an analgesic.

Tyrosine hydroxylase activity in the locus coeruleus of conscious rats: Differences with the anesthetized rats

Microdialysis was used to estimate in vivo tyrosine hydroxylase (TH) activity in locus coeruleus (LC) of conscious rats. An aromatic amino acid decarboxylase inhibitor, difluoromethyl-DOPA (DFMD), was perfused through the dialysis probe and the DOPA accumulating in dialysates was measured. A 3 h perfusion of a 6 × 10 −4 mol/ l DFMD solution was needed to obtain measurable levels of DOPA in LC dialy sates: a linear increase in DOPA concentrations was first observed and, then, a steady state. DOPA disappeared completely after inhibition of TH by α-methyl-p-tyrosine, showing that it reflects TH activity. The DFMD perfusion needed in this study was six times higher in concentration and three times longer in duration than the one required in a previous study we performed in LC of anesthetized rats. Furthermore, TH activity was found lower than the one we previously reported. We conclude that these differences could be explained by the effects of halothane anesthesia on DFMD and DOPA degradation and/or on LC neurons TH activity itself.

The mesolimbic dopamine-activating properties of ethanol are antagonized by mecamylamine

It has been suggested that ethanol may interact with the central nicotinic acetylcholine receptor, thus providing a basis for the often observed high consumption of both ethanol and nicotine. In the present in vivo microdialysis study, ethanol (2.5 g/kg) moderately increased dopamine overflow in the rat nucleus accumbens. The central nicotinic acetylcholine receptor antagonist mecamylamine totally counteracted this effect in a dose (1.0 mg/kg) that did not alter dopamine overflow per se. Ethanol also increased the overflow of dihydroxyphenylacetic acid and homovanillic acid, but this effect was not altered by mecamylamine (1.0 mg/kg). Furthermore, the ethanol-induced enhancement of 3,4-dihydroxyphenylalanine accumulation in the mesolimbic dopamine terminal area after NSD 1015 (an inhibitor of l-aromatic amino acid decarboxylase) was completely antagonized by mecamylamine in doses (3.0 and 6.0 mg/kg) that exerted no effects per se. Neither ethanol nor mecamylamine changed the catecholamine synthesis rate in the striatum or the cerebral cortex. These results provide further evidence that ethanol-induced activation of the mesolimbic dopamine system (increased dopamine synthesis and release) may be mediated via stimulation of central nicotinic acetylcholine receptors. It is suggested that antagonists of central nicotinic acetylcholine receptors may be useful in the treatment of alcoholism.

5-HT 1A autoreceptor-mediated effects of the amperozide congeners, FG5865 and FG5893, on rat brain 5-hydroxytryptamine neurochemistry in vivo

The two diphenylbutylpiperazinepyridinyl derivatives, FG5865 and FG5893, have a unique receptor binding profile in that they show very high and essentially equipotent affinities for both 5-HT 1A and 5-HT 2 receptors. The present report describes the acute effects of FG5865 and FG5893 on presynaptic 5-hydroxytryptamine (5-HT) neuronal function in the rat CNS, using established ex vivo and in vivo neurochemical techniques. Post-mortem measurements of tissue levels of 5-HT, its metabolite, 5-hydroxyindoleacetic acid (5-HIAA), and of the formation of 5-hydroxytryptophan (5-HTP; after inhibition of aromatic amino acid decarboxylase by NSD 1015) showed that FG5865 (0.1–20 mg/kg, s.c.) and FG5893 (0.1–20 mg/kg, s.c.) dose dependently decreased the synthesis and the metabolism/turnover of 5-HT - this to an extent comparable to the reference 5-HT 1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin. Reserpine (5 mg/kg, s.c.) pretreatment did not prevent the FG5893-induced decrease of 5-HT synthesis rate. In contrast, about 25–50 times higher doses of FG5865 were required to produce a comparable decrease in brain 5-HT synthesis in reserpinized vs. non-pretreated rats. In in vivo microdialysis experiments, both FG5865 (0.1–3.0 mg/kg, s.c) and FG5893 (0.3–1.0 mg/kg, s.c.) caused a marked and dose-dependent decrease of 5-HT release in the ventral hippocampus. Pretreatment with the 5-HT 1A receptor antagonist, (±)-pindolol (8 mg/kg, s.c.), abolished the FG5865 (0.3 mg/kg, s.c.)-induced reduction of 5-HT release, and (−)-pindolol (8 mg/kg, s.c.) similarly reversed the FG5893 (0.3 mg/kg, s.c.)-induced decrease. Local infusion of FG5865 into the ventral hippocampus (10 μM, 20-min pulse) resulted in a rapid and transient elevation of the 5-HT output, an effect that was independent of extracellular Ca 2+, FG5893, on the other hand, did not affect the 5-HT release upon local administration. The results demonstrate that FG5865 and FG5893 potently affect a range of neurochemical indices of rat brain 5-HT neuronal activity in vivo, in a way consistent with indirect (FG5865) and direct (FG5865 and FG5893) stimulation of the 5-HT 1A autoreceptors in the raphe nuclei.

Daily variations in in vivo tryptophan hydroxylation and in the contents of serotonin and 5-hydroxyindoleacetic acid in discrete brain areas of the rat.

The in vivo rate of brain tryptophan hydroxylation was determined through 5-hydroxytryptophan accumulation (5-HTPacc) following the administration of NSD 1015, a L-aromatic amino-acid decarboxylase inhibitor. This measurement was performed every 4 h throughout a 24 h hour period in 10 discrete brain areas of rats maintained on a regular 12 h/12 h light-dark cycle. The concentrations of 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) were also determined in untreated rats. Daily variations in 5-HTPacc were found in all the areas studied, the 5-HTPacc being higher during the dark period in most structures. These results strongly suggest that tryptophan hydroxylation is involved in the control of the 5-HT biosynthesis circadian rhythm. However, various patterns of 5-HT and 5-HIAA daily variations were observed, suggesting that the circadian factors affecting serotonin metabolism can be different among brain areas.

Activity of Tyrosine Hydroxylase in the Striatum of Newborn Piglets in Response to Hypocapnic Hypoxia

The effect of graded hypoxia induced by hyperventilation on the activity of tyrosine hydroxylase was measured in vivo by microdialysis. Microdialysis probes were inserted into the striatum of newborn piglets and perfused with medium containing 3-hydroxybenzylhydrazine, an inhibitor of L-aromatic amino acid decarboxylase. The level of 3,4-dihydroxyphenylalanine (DOPA) measured in the effluent dialysate was then an index of tyrosine hydroxylase activity. The oxygen pressure in the veins and capillaries of the cortex was measured, through a cranial window placed over the parietal cortex, by the phosphorescence lifetime of palladium-meso-tetra(4-carboxyphenyl)porphine added to the blood. After baseline measurements, PaCO2 was decreased from 38 torr (control value) to 19, 13, and 11 torr resulting in decreases in the cortical oxygen pressure from 40 +/- 6 torr to 26 +/- 3, 23 +/- 4, and 20 +/- 4 torr, respectively. Decrease in the oxygen pressure to 26 +/- 3 torr caused a statistically significant increase of 25-30% in the level of DOPA in the effluent perfusate. During the next step of increase in ventilator rate, when oxygen decreased only slightly, the level of DOPA remained at the higher level. Ventilation rates that lowered the oxygen pressure to below 20 torr, however, caused a progressive decrease in the level of DOPA. During recovery, the level of DOPA steadily increased, attaining 160% of control value after 1.5 h. When the oxygen pressure was decreased to 16 +/- 2 torr by a single increase in ventilator rate, the DOPA level decreased in the effluent to 15-20% below control.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of monoamine oxidase inhibitors on the acid metabolites of some trace amines and of dopamine in the rat striatum

The effects of the administration of selective and non-selective inhibitors of monoamine oxidase (MAO) on the concentrations of three trace acid metabolites [phenylacetic acid (PAA); m-hydroxyphenylacetic acid (mHPAA); and p-hydroxyphenylacetic acid (pHPAA)] and of an acid metabolite of dopamine [3,4-dihydroxyphenylacetic acid (DOPAC)] in the rat striatum were determined. Administration of brofaromine (1−100 mg/kg, s.c.) a type AMAO inhibitor, dose-dependently decreased DOPAC and mHPAA levels. pHPAA levels were decreased by 100 mg/kg brofaromine, but PAA levels were unaffected. Doses of deprenyl of less than 100 mg/kg, i.p., had no effect on any of the acids, while 100 mg/kg decreased DOPAC, mHPAA and pHPAA but not PAA levels. Clorgyline, pargyline and tranylcypromine treatment decreased the levels of DOPAC, mHPAA and pHPAA but not PAA. Administration of α-monofluoromethyldopa, an inhibitor of aromatic amino acid decarboxylase, decreased the levels of all four acids. It was concluded that deamination of the respective parent amine by type A MAO is primarily responsible for the synthesis of DOPAC and mHPAA, but that another pathway contributes to pHPAA synthesis. It appears that either PAA arises predominantly independently from the actions of MAO or that its removal via transport or further metabolism regulates its concentration.

Reduced Turnover of Dopamine and 5‐Hydroxytryptamine in Discrete Dopaminergic, Noradrenergic and Serotonergic Rat Brain Areas after Acutely Administered Medetomidine, a Selective α2‐Adrenoceptor Agonist

Monoamine metabolism and turnover were investigated in discrete dopaminergic, noradrenergic and serotonergic brain areas in the rat after acute administration of the selective alpha 2-adrenoceptor agonist, medetomidine. Medetomidine (3, 30 and 100 micrograms/kg subcutaneously) was given 90 min. before decapitation and discrete brain nuclei were punched from frozen brain slices for the analysis of concentrations of noradrenaline (NA), dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA). In a separate experiment, the accumulation of 3,4-dihydroxyphenylalanine (DOPA) and 5-hydroxytryptophan (5-HTP) was measured after inhibition of L-aromatic amino acid decarboxylase by NSD 1015: medetomidine (3, 10 and 100 micrograms/kg subcutaneously) was given 60 min. before NSD 1015 (100 mg/kg intraperitoneally), and the rates of DOPA and 5-HTP accumulation were determined over 30 min. Finally, the antagonistic effect of idazoxan (1 mg/kg subcutaneously), a selective alpha 2-adrenoceptor blocking agent, on the medetomidine-induced changes in monoamine metabolism was investigated. Medetomidine markedly decreased the metabolism and turnover of DA in the nucleus caudatus, but not in the nucleus accumbens or substantia nigra. In all dopaminergic areas, the turnover of 5-HT was markedly inhibited by medetomidine. These effects were significantly counteracted by idazoxan pretreatment demonstrating the alpha 2-receptor mediated action of medetomidine. The turnover of 5-HT was also reduced by medetomidine in the nucleus raphe dorsalis, the A1-C1 area, locus coeruleus, nucleus tractus solitarius and the A5 area. The accumulation of DOPA was markedly inhibited in the A1-C1 area, nucleus tractus solitarius and nucleus raphe dorsalis, but not in locus coeruleus.(ABSTRACT TRUNCATED AT 250 WORDS)

Stimulation of brain dopamine autoreceptors by remoxipride administration in reserpine-treated male rats.

Male Sprague-Dawley rats were treated subcutaneously with reserpine (5 mg kg-1, -18 h) and with the aromatic amino acid decarboxylase inhibitor, NSD-1015 (3-hydroxybenzylhydrazine) (100 mg kg-1 -30 min). Remoxipride 0.8, 3.2 or 12.8 mg kg-1 was administered subcutaneously at -50 min. Immediately following decapitation (0 h), the ventral striatum and the anterior neocortex were dissected. Dopa and 5-hydroxytryptophan accumulation in these brain areas were analysed by HPLC with electrochemical detection. Reserpine produced a marked increase in striatal and neocortical dopa accumulation, in comparison with glucose vehicle + NSD-1015-treated controls, and this increase was dose-dependently antagonized by remoxipride treatment. Thus, together with demonstrated dopamine receptor antagonist actions in intact animals, remoxipride behaves as a mixed dopamine receptor agonist-antagonist. Such properties could contribute to the favourable endocrine and extrapyramidal side effect profile of remoxipride as an antipsychotic agent.

Microdialysis monitoring of 3,4-dihydroxyphenylalanine accumulation after decarboxylase inhibition: a means to estimate in vivo changes in tyrosine hydroxylase activity of the rat locus ceruleus.

An on-line microdialysis approach was developed to estimate changes in tyrosine hydroxylase activity in the locus ceruleus noradrenergic neurons of anesthetized rats by measuring the 3,4-dihydroxyphenylalanine (DOPA) accumulation in the extracellular fluid during perfusion of an aromatic amino acid decarboxylase inhibitor through a dialysis probe. The aromatic amino acid decarboxylase inhibitor used was difluoromethyl-DOPA, which was shown to be more stable than NSD 1015 or Ro 4-4602 in the perfusion fluid. A 1-h perfusion of a 10(-4) mol/L of difluoromethyl-DOPA solution induced a linear increase in DOPA concentration in the locus ceruleus dialysates that achieved a steady state within 1 h. The identity of DOPA accumulated in dialysates during aromatic amino acid decarboxylase inhibition was confirmed by the disappearance of the chromatographic peak when DOPA formation was blocked by the administration of alpha-methyl-p-tyrosine. Systemic administration of the alpha 2-antagonist piperoxane before difluoromethyl-DOPA perfusion markedly increased the DOPA concentration during both the accumulation and the steady-state periods, showing that the present technique is a suitable in vivo approach to monitor changes in tyrosine hydroxylase activity occurring in the locus ceruleus neurons.

Effect of chlorpromazine, 5-hydroxytryptophan, and benserazide on plasma melatonin in the mudpuppy, Nectvrus maculosus

1. Melatonin (MEL) was measured by radioimmunoassay in adult Necturus maculosus treated with a melatonin precursor, 5-hydroxy- l-tryptophan (5HTP), a melatonin antimetabolite, chlorpromazine (CPZ), or an l-aromatic amino acid decarboxylase inhibitor, benserazide (BNS, Ro4-4062). MEL concentrations during photophase were significantly higher in animals receiving 5HTP (10, 20, 40 mg kg −1 ) than in saline-injected controls after 4 hr. 2. In animals receiving CPZ (2 or 20 mg kg −1) over a 3-day treatment schedule and sampled from 4 to 6 hr after injection, MEL was significantly higher than in controls whether sampled during midphotophase or midscotophase. Mean MEL in scotophase-sampled animals receiving 20 mg kg −1 was four times the concentration in midophotophase controls. 3. An ip dose of benserazide (50 or 80 mg kg −1) 3 hr after onset of photophase significantly reduced MEL compared with controls 3 hr after injection. The melatonin concentrations in animals receiving 80 mg kg −1 were only 4% of that in controls after 6 hr; MEL had recovered to 75% of control concentrations after 12 hr. 4. These experiments indicate that in Necturus: (1) MEL agonists can augment pathway enzyme activity during photophase; (2) a peripheral l-aromatic amino acid decarboxylase inhibitor operates under the same mechanism as in higher vertebrates; (3) chlorpromazine inhibition of MEL metabolism is effective day or night in increasing circulating MEL.

L-dopa-like regulatory actions of L-threo-3,4-dihydroxyphenylserine on the release of endogenous noradrenaline via presynaptic receptors in rat hypothalamic slices.

Effects of L-threo-3,4-dihydroxyphenylserine (L-threo-DOPS) on the spontaneous release and the stimulus(2 Hz)-evoked release of endogenous noradrenaline were studied in rat hypothalamic slices with functioning L-aromatic amino acid decarboxylase (AADC) and with AADC inhibition. In non-inhibited slices, spontaneous release was not modified by L-threo-DOPS at 1 pM-100 nM, tended to increase at 1-10 microM and increased at 100 microM. Noradrenaline tissue content slightly increased at 100 microM. Stimulated release was concentration-dependently facilitated at 1-1000 pM and tended to decrease gradually from a maximum at 10 nM-10 microM. Under AADC inhibition, spontaneous release concentration-dependently increased at 10-100 microM by 60% of the increase seen in slices without AADC inhibition. Increase in noradrenaline tissue content was abolished. L-threo-DOPS produced a triphasic pattern on stimulated release; concentration-dependent facilitation at 1-1000 pM similar to that seen in slices with functional AADC, no facilitation at 10-1000 nM, and a concentration-dependent increment at 10-100 microM. The facilitation at 1 nM was stereoselective and was antagonized by (-)-propranolol 10 nM, and no facilitation at 100 nM was restored to the maximum by yohimbine 10 nM, DG-5128 10 nM or S-sulpiride 1 nM. Furthermore, L-threo-DOPS (1-1000 pM)-induced facilitation was competitively antagonized by L-dopa methyl ester, a competitive antagonist for L-dopa, with a pA2 value of 13.6, whereas it was noncompetitively antagonized by (-)-propranolol.

The aromatic amino acid decarboxylase inhibitor, NSD-1015, increases release of dopamine: Response characteristics

Addition of the aromatic amino acid decarboxylase inhibitor, NSD-1015 (10 μM), to Krebs'-Ringer phosphate (KRP) superfusion medium, significantly increased the release of dopamine in vitro from superfused corpus striatum tissue fragments of male rats. A dose-dependent increase in release of dopamine was obtained in response to increasing concentrations of NSD-1015, with 1.0 μM being the minimally effective dose. In addition to the striatum, NSD-1015 also increased the release of dopamine from superfused hypothalamic tissue fragments. This capacity of NSD-1015 to increase release of dopamine was calcium-independent, appeared to be somewhat specific and could apparently increase the release of dopamine in vivo, as indicated by increases in the release of the metabolite of dopamine, DOPAC, under conditions of push-pull perfusion. Although the putative role of NSD-1015 is as an aromatic amino acid decarboxylase inhibitor, the present results demonstrate that, either as a result of this function and/or in addition to this role, NSD-1015 is a potent activator of the release of dopamine.