DOPA Concentration Research Articles

Serum Phenoloxidase Activity in the Hemolymph of Giant Freshwater Prawn Macrobrachium rosenbergii (De Man, 1879)

Melanization is one of the major immune responses in arthropods, particularly crustaceans. The prophenoloxidase (proPO) catalyzes the oxidation of mono or di or polyphenols, a reaction that is the key initial step of melanin formation. We attempted to study the immune response in M. rosenbergii by melanization reaction as produced by serum phenoloxidase (PO) activity. The substrate affinity of the PO enzyme was determined using different phenolic substrates and it was found that the mono, di and poly phenols were oxidized hence the enzyme was characterized as catecholoxidase and l-3,4-dihydroxyphenylalanine (L–DOPA) showed the highest substrate affinity to the enzyme. The biochemical parameters that determined optimum enzyme activity were found such as 1 mM L–DOPA concentration showing at the absorbance of 470 nm, 10 mM Tris–HCl buffer pH 7.5 (brown colour formation), temperature 40 ° C and 10 min incubation. Kinetic characteristics of serum from the prawn were also determined. Determination of optimal conditions of PO activity in the serum has also been attempted. We also identified the serum having a potent PO activity was inhibited by phenylthiourea (PTU) and activation of such as trypsin, sodium dodecyl sulphate (SDS) and laminarin in the freshwater prawn M. rosenbergii.

Lightweight and flexible silicone rubber foam with dopamine grafted multi-walled carbon nanotubes and silver nanoparticles using supercritical foaming technology: Its preparation and electromagnetic interference shielding performance

Herein, dopamine is grafted onto multi-walled carbon nanotubes (MWCNTs), and then grafted dopamine is used as a chemical activation point to initiate the in-situ synthesis of silver nanoparticles (Ag-NPs). The foam technology with supercritical N2 (Sc-N2) gas was used to prepare a lightweight and flexible silicone rubber foam material with efficient electromagnetic shielding performance. The prepared material has uniform cells and a density of only 0.3 g/cm3. The electrical conductivity can be achieved as high as 5.69 S/m. At the same time, the optimal modified conductive filler MWCNTs conditions are determined: DOPA concentration is 2 g/L, deposition time is 18 h, formaldehyde concentration is 5 g/L and AgNO3 concentration is 16 g/L. From the perspective of electromagnetic interference shielding performance (EMI SE), the value of prepared silicone rubber foam with 15 parts of MWCNT-PDA-Ag (VMPA15) can reach 36 dB, far exceeding the commercial requirement (20 dB). The EMI shielding performance is also improved with the increased thickness of the sample. When the sample thickness is 10 mm, the EMI SE value can be as high as 52 dB; more importantly, this type of VMPA foam has a shielding mechanism based on absorption, which is the most effective for solving the problem of secondary magnetic pollution caused by electromagnetic wave reflection.

Pathogenetic treatment of patients with hypertension depending on the level of sympathоadrenal system activity

Purpose of the study. Recently, new opportunities to optimize clinical efficacy and tolerability of treatment of patients with essential hypertension appeared. Determination of the indicators of increased activity of the sympathetic nervous system (SNS) in practice could contribute to differentiated treatment of patients with hypertension. Comparative estimation of influence of complex treatment with bisoprolol or amlodipine on the SNS activity parameters in patients with arterial hypertension (AH) of II stage.
 Materials and methods. 121 patients (men and women) with documented II stage AH were examined clinically. Concentrations of adrenaline, noradrenaline, dopamine and DOPA in urine were evaluated by spectrochromatography, endotheliumdependent vasodilation was studied using complex rheographic ReoCom Professional (HAI-Medicom, Ukraine). Patients were divided into 2 groups: if concentration of noradrenalin in urine exceeds healthy subjects levels more than on 20% – patients were added to basic treatment (enalapril, atorvastatin, aspirin) bisoprolol, if concentration of noradrenalin in urine exceeds healthy subjects levels less than on 20% – we added amlodipine. Statistic analysis was done on Apache OpenOffice (version 4.1) and PSPP (version 0.7.9).
 Results. The study showed that after treatment there was significant decrease of adrenalin, noradrenalin in 1st group . Increase of DOPA levels was higher in 1st group (Δ% = +46,5 [30,0; 55,4]% vs Δ% = +8,8 [–2,1 ; 16,0]%, (p < 0,05).). Trends in daily urinary excretion of adrenaline (–44,8%) and norepinephrine (–33,7%) showed the impact of effective Δ-blocker on the activity of pressor systems. DOPA excretion was increased significantly in both groups which can indirectly indicate hemodynamic load reduction in hypertensive patients. When choosing the individual antihypertensive treatment in patients with arterial hypertension, if excretion of noradrenalin is increased it is rationally to add adjuvant therapy with β-blockers.
 Conclusion. The results of our research can be applied into the practice of general practitioner as recommendations for individual selection of antihypertensive treatment depending on the activity of sympathetic nervous system.

Flow-Injection Amperometric Determination of DOPA and Tyrosine at a Dual Electrode Modified with the Gold–Cobalt Binary System

The gold–cobalt binary system, electrodeposited on the surface of a screen-printed electrode, exhibits catalytic activity in the electrooxidation of DOPA and tyrosine. The catalytic effect is shown in a multiple increase in current compared with the oxidation current of the modifier and a decrease in the oxidation overvoltage of organic compounds. Methods for the flow-injection amperometric determination of DOPA and tyrosine at the proposed modified electrode are developed. The simultaneous amperometric determination of DOPA and tyrosine at a dual screen-printed electrode modified with an Au–Co binary system the under conditions of flow-injection analysis is demonstrated. The linear dependence of the analytical signal on the concentration of DOPA and tyrosine is observed in the ranges from 1 × 10–9 to 1 × 10–4 M and from 5 × 10–8 to 5 × 10–4 M, respectively.

Electrochemical marker for metastatic malignant melanoma based on the determination of l-dopa/l-tyrosine ratio

In this paper, we have proposed an electrochemical marker for the metastatic malignant melanoma by determining 3,4-dihydroxy-l-phenylalanine (l-dopa or DOPA) and l-tyrosine (Tyr) ratio using the electropolymerized film of 3-amino-5-mercapto-1,2,4-triazole on glassy carbon (p-AMTa) electrode. Earlier studies suggested that DOPA/Tyr ratio can be used to determine the aggressiveness of malignant melanoma. Based on this report, we attempted to determine the concentrations of DOPA and Tyr simultaneously using p-AMTa electrode by electrochemical method. The voltammetric signals of DOPA and Tyr were not resolved when we use bare GC electrode. However, p-AMTa electrode not only separates the voltammetric signals of DOPA and Tyr but also enhanced their oxidation currents. Using the p-AMTa film, we have successfully determined DOPA and Tyr simultaneously for the first time. To demonstrate the clinical application of the present method, we have successfully determined the concentrations of DOPA and Tyr in human blood plasma samples (n=4) and estimated DOPA/Tyr ratio. The ratio was found within the prescribed limit, indicating that the blood plasma samples are free from melanoma disease.

Determinants of buildup of the toxic dopamine metabolite DOPAL in Parkinson's disease

Intra-neuronal metabolism of dopamine (DA) begins with production of 3,4-dihydroxyphenylacetaldehyde (DOPAL),which is toxic. According to the 'catecholaldehyde hypothesis', DOPAL destroys nigrostriatal DA terminals and contributes to the profound putamen DA deficiency that characterizes Parkinson’s disease (PD). We tested the feasibility of using post-mortem patterns of putamen tissue catechols to examine contributions of altered activities of the type 2 vesicular monoamine transporter (VMAT2) and aldehyde dehydrogenase(ALDH) to the increased DOPAL levels found in PD. Theoretically, the DA : DOPA concentration ratio indicates vesicular uptake, and the 3,4-dihydroxyphenylacetic acid: DOPAL ratio indicates ALDH activity. We validated these indices in transgenic mice with very low vesicular uptake VMAT2-Lo) or with knockouts of the genes encoding ALDH1A1 and ALDH2 (ALDH1A1,2 KO), applied these indices in PD putamen, and estimated the percent decreases in vesicular uptake and ALDH activity in PD. VMAT2-Lo mice had markedly decreased DA:DOPA (50 vs. 1377, p < 0.0001),and ALDH1A1,2 KO mice had decreased 3,4-dihydroxyphenylacetic acid:DOPAL (1.0 vs. 11.2, p < 0.0001). In PD putamen, vesicular uptake was estimated to be decreased by 89% and ALDH activity by 70%. Elevated DOPAL levels in PD putamen reflect a combination of decreased vesicular uptake of cytosolic DA and decreased DOPAL detoxification by ALDH.

An increase in renal dopamine does not stimulate natriuresis after fava bean ingestion

Fava beans (Vicia faba) contain dihydroxyphenylalanine (dopa), and their ingestion may increase dopamine stores. Renal dopamine regulates blood pressure and blood volume via a natriuretic effect. The objective was to determine the relation between dietary fava beans, plasma and urinary catechols, and urinary sodium excretion in 13 healthy volunteers. Catechol and sodium data were compared by using a longitudinal design in which all participants consumed a fixed-sodium study diet on day 1 and the fixed-sodium diet plus fava beans on day 2. Blood was sampled at 1, 2, 4, and 6 h after a meal, and 3 consecutive 4-h urine samples were collected. Mean (±SD) plasma dopa was significantly greater 1 h after fava bean consumption (11,670 ± 5440 compared with 1705 ± 530 pg/mL; P = 0.001) and remained elevated at 6 h. Plasma dopamine increased nearly 15-fold during this period. Fava bean consumption also increased urinary dopamine excretion to 306 ± 116, 360 ± 235, and 159 ± 111 μg/4-h urine sample compared with 45 ± 21, 54 ± 29, and 44 ± 17 μg in the 3 consecutive 4-h samples after the control diet (P ≤ 0.005). These substantial increases in plasma and urinary dopa and dopamine were unexpectedly associated with decreased urinary sodium. The failure of fava bean consumption to provoke natriuresis may indicate that dopa concentrations in commercially available beans do not raise renal dopamine sufficiently to stimulate sodium excretion, at least when beans are added to a moderate-sodium diet in healthy volunteers. This trial was registered at clinicaltrials.gov as NCT01064739.

Program Summary

Objectives: 6-[18F]Fluoro-L-DOPA (FDOPA) is used to study the presynaptic dopamine synthesis in vivo using PET [1]. Major indications for this tracer are determination of the disease stage in Parkinson as well as detection of various neuroendocrine tumors and their metastases. Prior to set up a Good Manufacturing Practice (GMP) compliant production, the quality control of FDOPA needs to be validated. The radiopharmaceutical needs to meet the specification as described in the European Pharmacopoeia 7.0 Methods: An Ultra Performance Liquid Chromatography UPLC® with an online radioactivity detector was used to measure the concentrations of FDOPA and impurities. For identification of the parent compound and its impurities an UPLC® coupled to a Xevo® G2 QTof mass spectrometer was used. The different components were separated on an Waters ACQUITY UPLC® HSS T3 1.8 μm, 3.0 x 50 mm analytical column. The eluent is 0.05 M sodium dihydrogen phosphate buffer pH 2.5 with an isocratic flow of 0.8 ml/min. Total run time is 3 minutes. Several calibration samples of FDOPA were measured to determine linearity, reproducibility, stability, LOQ, LOD and the resolution. Furthermore the linearity of the Berthold flowstar LB 513 online radioactivity detector was measured. For identification of the parent compound and impurities, a sample was taken directly after the [18F]FDOPA synthesis. The measurements with the mass spectrometer were performed in negative resolution mode with an electrospray source in a range from 50 to 1200 Da. The masses of interest of FDOPA, DOPA, 6-hydroxy- DOPA and FDOPA-quinone (see fig. 1) are respectively 214.0516, 196.0610, 212.0559 and 212.0359 Da. (Figure presented) Results: The analyses using UPLC® in combination with online radioactivity detector were validated. FDOPA and side products DOPA and FDOPA-quinone were detected and identified in the production batches. The concentrations were 1040, 0.1 and 61.5 μg/mL (n = 19), respectively. The samples were measured over a period time of 9 weeks after production and showed a clear increase in the concentration of FDOPA- quinone over time. The DOPA concentration decreasing over this period. In all the samples the concentration of 6-hydroxy- DOPA was below the detection limit and therefore it was not possible to identify this side product with UPLC® -MS. The main impurity was FDOPA-quinone, which is not described in the European Pharmacopoeia 7.0. Conclusions: The quality control of FDOPA using UPLC® -RA was validated for GMP productions. The UPLC® method is reproducible and the retention times of the desired compounds are very short. This is of advantage for rapid releasing the production batch for human application.

Gold mesoparticles with precisely controlled surface topographies for single-particle surface-enhanced Raman spectroscopy

Understanding of the influence of nanotextured surface topographies on surface-enhanced Raman scattering (SERS) is still rather limited, even after intensive research in recent years; this is mainly due to the absence of a synthesis system to control the surface morphologies on metal meso-/nanoparticles. Here, a facile particle-mediated aggregation protocol was reported to synthesize a series of uniform gold mesoparticles with a nanotextured rough surface. The surface topographies can be precisely controlled by varying the temperatures and concentrations of dopa and Au ions. For example, the tips can be well changed from half-sphere to spike, and then to blade, and the size of the tips also can be tuned, such that the thickness of the blade-like tips can be artificially controlled from 9 to 14 nm. Single-particle SERS analysis reveals that the uniform Au mesoparticles with highly nanotextured topography have enhancement factors greater than 1.0 × 108 with narrow distribution. The effect of the precisely controlled mesoparticle topographies on SERS properties was deeply studied using Raman analysis and finite-difference time-domain (FDTD) simulation. The result indicated that a strong electric field will be induced at the tips and slits on the flower-like mesoparticle surface by the confined light in the cavities between the petals. As such, the current protocol opens up avenues for the fabrication of structurally reproducible mesosuperstructure-based SERS sensors.

The Contribution of DOPA to Substrate–Peptide Adhesion and Internal Cohesion of Mussel‐Inspired Synthetic Peptide Films

Mussels use a variety of 3, 4-dihydroxyphenyl-l-alanine (DOPA) rich proteins specifically tailored to adhering to wet surfaces. Synthetic polypeptide analogues of adhesive mussel foot proteins (specifically mfp-3) are used to study the role of DOPA in adhesion. The mussel-inspired peptide is a random copolymer of DOPA and N(5) -(2-hydroxyethyl)-l-glutamine synthesized with DOPA concentrations of 0-27 mol% and molecular weights of 5.9-7.1 kDa. Thin films (3-5 nm thick) of the mussel-inspired peptide are used in the surface forces apparatus (SFA) to measure the force-distance profiles and adhesion and cohesion energies of the films in an acetate buffer. The adhesion energies of the mussel-inspired peptide films to mica and TiO(2) surfaces increase with DOPA concentration. The adhesion energy to mica is 0.09 μJ m(-2) mol(DOPA) (-1) and does not depend on contact time or load. The adhesion energy to TiO(2) is 0.29 μJ m(-2) mol(DOPA) (-1) for short contact times and increases to 0.51 μJ m(-2) mol(DOPA) (-1) for contact times >60 min in a way suggestive of a phase transition within the film. Oxidation of DOPA to the quinone form, either by addition of periodate or by increasing the pH, increases the thickness and reduces the cohesion of the films. Adding thiol containing polymers between the oxidized films recovers some of the cohesion strength. Comparison of the mussel-inspired peptide films to previous studies on mfp-3 thin films show that the strong adhesion and cohesion in mfp-3 films can be attributed to DOPA groups favorably oriented within or at the interface of these films.

N-(3,5-Dihydroxybenzoyl)-6-hydroxytryptamine as a Novel Human Tyrosinase Inhibitor That Inactivates the Enzyme in Cooperation with L-3,4-Dihydroxyphenylalanine

N-(3,5-Dihydroxybenzoyl)-6-hydroxytryptamine (2) was a novel inhibitor of L-3,4-dihydroxyphenylalanine (DOPA) oxidase activity of human HMV-II melanoma tyrosinase. The IC₅₀ values for 2 and three reference compounds, N-(3,5-dihydroxybenzoyl)serotonin, 6-hydroxyindole, and kojic acid, were 9.1, 842, 22, and 310 µM, respectively, indicating that the 6-hydroxyindole moiety was more effective than 5-hydroxyindole as the pharmacophore of polyphenolic tyrosinase inhibitors and that the inhibitory activity of 6-hydroxyindole was strengthened by the link with a resorcinol group. Furthermore, compound 2 exhibited a unique property of inactivating the human tyrosinase in the presence of low concentrations of DOPA. This inactivation was attenuated by high concentrations of DOPA and for the most part was irreversible as confirmed by activity stain in native polyacrylamide gel electrophoresis and by removal of 2 and DOPA using gel permeation chromatography. Tyrosinase is the enzyme that oxidizes tyrosine to DOPA and further oxidizes DOPA to the melanin precursor dopaquinone. A compound such as 2 that inactivates the enzyme in the presence of a small amount of DOPA is therefore attractive as a new type of tyrosinase inhibitor. Unfortunately, 2 hardly suppressed the melanogenesis in melanoma cell culture. However, the above strong inhibitory activity and the unique property in the combination with DOPA suggest that this compound is a useful lead in designing new antimelanogenic agents.

Amine neuromediators, their precursors, and oxidation products in the culture of Escherichia coli K-12

Growth dynamics of the synthesis of monoamine neuromediators serotonin, norepinephrine, and dopamine in Escherichia coli K-12 was investigated for the first time using high performance liquid chromatography with electrodetection. Maximum (micromolar) concentrations of these compounds were detected in E. coli cells during the early growth phases; their intracellular content decreases after the transition to late growth phases. E. coli biomass contains the substances DOPA and 5-hydroxytryptamine that serve in animal cells as neuromediator precursors and the products of their oxidative deamination. Presumably, the biosynthesis and degradation of monoamine neuromediators in bacterial cells involves enzyme systems analogous to those typical of animals. The culture fluid of E. coli contains micromolar concentrations of DOPA and nanomolar of serotonin, dopamine, and norepinephrine during the late growth phase. These concentrations are sufficient for animal/human receptors to bind them. This article deals with the potential biotechnological applications of the data obtained.

Complexities of cuticular pigmentation in insects

Complexities of cuticular pigmentation in insects

Stability of collagen in the presence of 3,4-dihydroxyphenylalanine (DOPA)

Many cross-linking agents for collagen are available with varying levels of toxicity and some are in use in biomedical implants of collagen. L-DOPA (3,4-dihydroxyphenylalanine), a neurotransmitter, is a naturally present compound in the living system and is the target in therapeutic strategy of Parkinson’s disease. This work reports the effect of the neurotransmitter DOPA on the stability of collagen solution using circular dichroism (CD), fluorescence spectroscopy, melting and shrinkage temperature. Collagen solution treated with various concentrations of DOPA ranging from 10 −2 to 10 −5 M was analyzed using fluorescence and CD spectra. When collagen was treated with DOPA, the intensity of emission was found to increase indicating the possibility of interaction of DOPA with collagen and maximum emission intensity was observed between 10 −3 and 10 −4 M for L-DOPA and DL-DOPA, respectively. CD studies show possible aggregation of collagen even in the presence of low concentrations of DOPA. The shrinkage temperature of DOPA treated collagen fibres was experimentally determined to be 69 ± 1 °C. The melting temperature of DOPA cross linked collagen solution also exhibited a significant increase from 35 to 40 °C (±0.1) ( P < 0.05). The experimental results suggest that the optimum concentration for cross linking collagen with DOPA ranges between 10 −3 and 10 −4 M. Thus, DOPA may be a useful stabilizing agent for collagen for biomedical applications.

In vitro interaction of L-Dopa with bacterial adhesins of Helicobacter pylori: An explanation for clinicial differences in bioavailability?

Objectives Recent investigations on the pharmacokinetics of levodopa ( L -dopa) indicated that the presence of Helicobacter pylori in patients with Parkinson's disease, orally treated with L -dopa, influences the absorption of this compound, which consequently leads to decreased plasma levels. Therefore this work aims to study a potential in-vitro interaction of L -dopa with H. pylori and its surface adhesins. Methods Solutions containing L -dopa of different concentrations were incubated with H. pylori at different bacterial densities and time intervals. Free L -dopa was quantified from the incubation supernatants by HPLC. A flow cytometric assay with fluorescence labelled H. pylori was used to investigate the influence of L -dopa on the bacterial adhesion of H. pylori: FITC-labelled bacteria were pre-incubated with L -dopa, followed by incubation with gastric epithelial cells (AGS cells) and FACS quantification of adhering bacteria. Key findings Evaluation of time- and concentration-dependent incubation experiments indicated a significant decrease in L -dopa concentrations when coming into contact with H. pylori. The reduction in L -dopa concentrations was determined as 47 to 12%, referred to the initial starting concentration, with time-dependency and dependency of the H. pylori density. FITC-labelled H. pylori, pre-incubated with differing L -dopa concentrations, were shown to have a significantly reduced bacterial adhesion to AGS cells, with a maximum reduction of 22 ± 9%. These results demonstrate a direct interaction of L -dopa with the outer membrane proteins of H. pylori responsible for the adhesion to gastric epithelial cells. By this interaction the unbound L -dopa concentration in bacterial suspension was strongly reduced. Conclusions This study suggests a potential in-vitro interaction of L -dopa with H. pylori adhesins, confirming the clinical changes found in pharmacokinetics of L -dopa therapy by H. pylori-positive patients with Parkinson's disease.

Biomarkers to detect central dopamine deficiency and distinguish Parkinson disease from multiple system atrophy

Objective Biomarkers are increasingly important to diagnose and test treatments of neurodegenerative diseases such as Parkinson disease (PD). This study compared neuroimaging, neurochemical, and olfactory potential biomarkers to detect central dopamine (DA) deficiency and distinguish PD from multiple system atrophy (MSA). Methods In 77 PD, 57 MSA, and 87 control subjects, radioactivity concentrations in the putamen (PUT), caudate (CAU), occipital cortex (OCC), and substantia nigra (SN) were measured 2 h after 6-[ 18F]fluorodopa injection, septal myocardial radioactivity measured 8 min after 6-[ 18F]fluorodopamine injection, CSF and plasma catechols assayed, or olfaction tested (University of Pennsylvania Smell Identification Test (UPSIT)). Receiver operating characteristic curves were constructed, showing test sensitivities at given specificities. Results PUT:OCC, CAU:OCC, and SN:OCC ratios of 6-[ 18F]fluorodopa-derived radioactivity were similarly low in PD and MSA ( p < 0.0001, p < 0.0001, p = 0.003 compared to controls), as were CSF dihydroxyphenylacetic acid (DOPAC) and DOPA concentrations ( p < 0.0001, each). PUT:SN and PUT:CAU ratios were lower in PD than in MSA ( p = 0.004; p = 0.005). CSF DOPAC correlated positively with PUT:OCC ratios ( r = 0.61, p < 0.0001). Myocardial 6-[ 18F]fluorodopamine-derived radioactivity distinguished PD from MSA (83% sensitivity at 80% specificity, 100% sensitivity among patients with neurogenic orthostatic hypotension). Only PD patients were anosmic; only MSA patients had normal olfaction (61% sensitivity at 80% specificity). Conclusions PD and MSA feature low PUT:OCC ratios of 6-[ 18F]fluorodopa-derived radioactivity and low CSF DOPAC and DOPA concentrations, cross-validating the neuroimaging and neurochemical approaches but not distinguishing the diseases. PUT:SN and PUT:CAU ratios of 6-[ 18F]fluorodopa-derived radioactivity, cardiac 6-[ 18F]fluorodopamine-derived radioactivity, and olfactory testing separate PD from MSA.

Allopregnanolone modulates striatal dopamingergic activity of rats under different gonadal hormones conditions

Objectives: Progesterone modulates dopamine (DA) release in corpus striatum. Our objective was to evaluate the effect of the i.c.v injection of the neurosteroid allopregnanolone (ALL), a progesterone metabolite on dopaminergic activity in the corpus striatum of rats under different gonadal hormonal conditions.Methods: We have measured the concentrations of DOPA, DA and DOPAC (main metabolite of DA) in the corpus striatum in estrus and diestrus rats and in ovariectomized rats without hormonal replacement (OVX group) and primed with estrogen and progesterone (OVXi group). Additionally, we have used the aromatic acid decarboxylase inhibitor NSD in order to evaluate the function of tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine synthesis.Results: ALL significantly decreased the striatal concentrations of both DA and DOPAC in the estrus. On the other hand, ALL increased significantly the levels of DA in the OVXi group. The DOPA accumulation in OVXi after NSD treatment in the ALL-treated groups was greater than in the vehicle group. However, the estrus group did not modify the DOPA accumulation after NSD injection.Discussion: Our results suggest that ALL could modulate the dopaminergic transmission in the corpus striatum by causing changes in the activity of TH and/or in the pre- and post-synaptic dopaminergic terminals in the corpus striatum. This neurosteroidal mechanism could be a new kind of neurotransmitter systems modulation accomplished on TH activity itself and/or on the second messengers not related to ionic channels. Additionally, our results reinforce the idea of a close relationship between the fast non-genomic mechanism of ALL and the genomic actions of estrogen and progesterone.

Extracellular striatal concentrations of endogenous 3,4-dihydroxyphenylalanine in the absence of a decarboxylase inhibitor: a dynamic index of dopamine synthesis in vivo.

Basal levels of endogenous 3,4-dihydroxyphenylalanine (DOPA) were detected by HPLC coupled with coulometric detection in dialysates from freely moving rats implanted 48-72 h earlier with transversal dialysis fibers in the dorsal caudate. Because decarboxylase inhibitor is absent in the Ringer's solution, this method allows monitoring of basal output of dopamine (DA) and 3,4-dihydroxyphenylacetic acid, as well as DOPA. Extracellular DOPA concentrations were reduced by the tyrosine hydroxylase inhibitor alpha-methylparatyrosine (200 mg/kg, i.p.) and by the dopaminergic agonist apomorphine (0.25 mg/kg, s.c.). The dopaminergic antagonist haloperidol (0.2 mg/kg, s.c.) stimulated DOPA output by about 60% over basal values. Gamma-Butyrolactone, at doses of 700 mg/kg, i.p., which are known to block dopaminergic neuronal firing and which reduce DA release, stimulated DOPA output maximally by 130% over basal values. Tetrodotoxin, which blocks DA release by blocking voltage-dependent Na+ channels, increased DOPA output maximally by 100% over basal values. The results indicate that basal DOPA can be detected and monitored in the extracellular fluid of the caudate of freely moving rats by transcerebral dialysis and can be taken as a dynamic index of DA synthesis in pharmacological conditions.

Effect of 3,4-dihydroxyphenylalanine on Cu2+-induced Inactivation of HDL-associated Paraoxonase1 and Oxidation of HDL; Inactivation of Paraoxonase1 Activity Independent of HDL Lipid Oxidation

Paraoxonase1 (PON1), one of HDL-asssociated antioxidant proteins, is known to be sensitive to oxidative stress. Here, the effect of endogenous reducing compounds on Cu2+-mediated inactivation of PON1 was examined. Cu2+-mediated inactivation of PON1 was enhanced remarkably by catecholamines, but not by uric acid or homocysteine. Furthermore, catecholamines such as 3,4-dihydroxyphenylalanine (DOPA), dopamine or norepinephrine were more effective than caffeic acid or pyrocatechol in promoting Cu2+-mediated inactivation of PON1, suggesting the importance of dihydroxybenzene group as well as amino group. DOPA at relatively low concentrations showed a concentration-dependent inactivation of PON1 in a concert with Cu2+, but not Fe2+. The DOPA/Cu2+-induced inactivation of PON1 was prevented by catalase, but not hydroxyl radical scavengers, consistent with Cu2+-catalyzed oxidation. A similar result was also observed when HDL-associated PON1 (HDL-PON1) was exposed to DOPA/Cu2+. Separately, it was found that DOPA at low concentrations (1-6 μM) acted as a pro-oxidant by enhancing Cu2+-induced oxidation of HDL, while it exhibited an antioxidant action at ≥10 μM. In addition, Cu2+-oxidized HDL lost the antioxidant action against LDL oxidation. Meanwhile, the role of DOPA/Cu2+-oxidized HDL differed according to DOPA concentration; HDL oxidized with Cu2+ in the presence of DOPA (60 or 120 μM) maintained antioxidant activity of native HDL, in contrast to an adverse effect of DOPA at 3 or 6 μM. These data indicate that DOPA at micromolar level may act as a pro-oxidant in Cu2+-induced inactivation of PON1 as well as oxidation of HDL. Also, it is proposed that the oxidative inactivation of HDL-PON1 is independent of HDL oxidation.

Role of tyrosine, DOPA and decarboxylase enzymes in the synthesis of monoamines in the brain of the locust

The metabolic transformation of tyrosine (TYR) by the decarboxylase and hydroxylase enzymes was investigated in the central nervous system of the locust, Locusta migratoria. It has been demonstrated that the key amino acids, 3,4-dihydroxyphenylalanine (DOPA), 5-hydroxytryptophan (5HTP) and tyrosine are decarboxylated in all part of central nervous system. DOPA and 5HTP decarboxylase activities show parallel changes in the different ganglia, but the rank order of the activity of TYR decarboxylase is different. Enzyme purification has revealed that the molecular weights of TYR decarboxylase and DOPA/5HTP decarboxylase are 370,000 and 112,000, respectively. The decarboxylation of DOPA by DOPA/5HTP decarboxylase is stimulated, whereas the decarboxylation of DOPA by TYR decarboxylase is inhibited in the presence of the cofactor pyridoxal-5′-phosphate. TYR hydroxylase could not be detected and 3 H -TYR is found to be metabolised to tyramine (TA), but not to DOPA. The haemolymph contains a significant concentration of DOPA (120 pmol/100 μl haemolymph), and the ganglia incorporates DOPA from the haemolymph by a high affinity uptake process ( K M=12 μM and V max=24 pmol per ganglion/10 min). Our results suggest that no tyrosine hydroxylase is present in the locust CNS and the DOPA uptake into the ganglia by a high affinity uptake process as well as the DOPA decarboxylase enzyme may be responsible for the regulation of the ganglionic dopamine (DA) level. Two types of decarboxylases exist, one of them decarboxylating DOPA and 5HTP (DOPA/5HTP decarboxylase), other decarboxylating TYR (TYR decarboxylase). The DOPA/5HTP decarboxylase enzyme present in the insect brain may correspond to the 5HTP/DOPA decarboxylase in vertebrate brain, whereas TYR decarboxylase is characteristic only for the insect brain.