Dietary calcium supplementation and dopamine-β-hydroxylase in spontaneously hypertensive rats

Spontaneously hypertensive 4-week-old male rats were fed, before and after the onset of hypertension, with either commercial chow (control) or commercial chow combined with different forms of milk proteins with or without calcium supplementation. After 40 weeks, rats were still hypertensive, and dopamine-β-hydroxylase enzyme activity measured simultaneously in serum and adrenal was found to be higher than in the controls. The enzyme activity in rats fed diets with milk proteins was increased significantly in both serum and adrenal compared with the control, and such enhancement was significantly higher than that observed in animals fed the commercial diet supplemented with calcium (1.2%), suggesting that dietary calcium intake associated with dietary protein of high digestibility, such as casein, potentiates the endogenous mechanisms regulating the homeostasis of calcium more than calcium supplementation itself. Moreover, the selective and additive effect of diets supplemented with milk proteins and calcium on adrenal enzyme activity clearly suggests a relationship between cardiovascular diseases involving the genesis of hypertension and stress mechanisms through the hypothalamo-pituitary adreno-sympathetic axis.     

Serum dopamine-β-hydroxylase in psychiatric patients and normals. Effect of d-amphetamine and haloperidol

Serum dopamine--hydroxylase activity was estimated in groups of normals and of psychiatric patients, using a thin layer radiochromatographic method. The percentage of patients with schizophrenic and with depressive symptomatology was higher in the population with high enzyme activities. In addition, d-amphetamine given to normals caused an increase in the serum activity while haloperidol caused the opposite effect. The activity in serum is interpreted as a loss in the enzyme from the place it acts physiologically, with possible influence on the noradrenaline synthesis rate.     

Plasma dopamine-β-hydroxylase activity and the pressor effect of dopamine infusion in man

Summary In order to study the function of dopamine--hydroxylase (DBH) in human plasma, dopamine, its natural substrate, was infused intravenously in 22 healthy volunteers. Their plasma DBH activities showed great interindividual variations (31–301 units/ml). The infusion rates of dopamine required to increase systolic blood pressure (BP) by 30 mm Hg differed considerably between the subjects, and ranged from 3,0 to 11,6 µg/kg/min. No correlation could be shown between the various dopamine doses and individual plasma levels of DBH. It was concluded, therefore, that plasma DBH in the blood stream was enzymatically inactive. Experiments with human plasma DBH in vitro also support this interpretation. Consequently, interindividual differences in the effects on BP during dopamine infusion cannot be due to pressor effects of noradrenaline synthesized by plasma DBH.The study was supported by the Deutsche Forschungsgemeinschaft     

Absence of a relationship between sympathetic neuronal activity and turnover of serum dopamine-β-hydroxylase

Summary The effects of pharmacological alteration of adrenergic transmission on the rate of entrance of dopamine--hydroxylase (DBH) into the circulation were assessed in rats by an immunological method in which the kinetics of recovery of serum DBH activity were measured after depletion of the enzyme by treatment with anti-rat DBH antiserum. Neither -receptor blockade with phenoxybenzamine nor ganglionic blockade with clorisondamine altered the rate by which DBH enters the bloodstream although both treatments markedly altered serum catecholamine levels. Prolonged treatment of newborn rats with guanethidine produced a severe peripheral sympathectomy but only a moderate decrease (30%) in serum DBH levels. In the sympathectomized rats, the rate of entrance of DBH into the circulation was significantly reduced whereas the half-life and rate of degradation of the enzyme was unchanged. These results indicate that the major portion of serum DBH does not enter the circulation by means of exocytotic release of the soluble enzyme.     

Human circulating dopamine-Β-hydroxylase and epilepsy

The activity of circulatory dopamine--hydroxylase (DBH) in humans is shown to be lower in some epileptic subjects than in normal subjects. The activity of the enzymes was found to be dramatically low in subjects who experienced an epileptic seizure 24 hrs before DBH activity was determined. The activity varied through the course of epileptic seizures induced by a convulsant drug and these variations might be due to the en masse changes of the sympathetic nervous system.Abbreviations DBH Dopamine--hydroxylase - Bemegride 4-methyl-4-ethyl-2,6-dioxopiperidine This work is a part of the Doctorat d'Université Thésis of M.T. M.-P., recipient of the Juan March Fundation.D.A. is Attaché de Recherche at the INSERM.Service d'Exploration fonctionelle du Système Nerveux.     

Determination of dopamine-β-hydroxylase in cerebrospinal fluid by high-performance liquid chromatography with electrochemical detection

An improved method is described for the measurement of dopamine-β-hydroxylase (DβH) activity in cerebrospinal fluid, which is based on an incubation with dopamine at a saturated substrate concentration and quantitation of the reaction product noradrenaline, by high-performance liquid chromatography with electrochemical detection using 3,4-dihydroxynorephedrine as internal standard. Sample workup consists in an ion pair extraction to isolate the catecholamines from the rather complex incubation medium, a cation ion exchange to eliminate the bulk amount of dopamine, and alumina adsorption to concentrate the sample prior to high-performance liquid chromatography. The methodology was used to evaluate some of the characteristics of DβH in cerebrospinal fluid and the stability of the enzyme. The procedure was also employed to determine the change in the DβH following drug administration. Intravenously administered yohimbine caused an increase in DβH activity in cerebrospinal fluid of rabbits as expected from its known α₂-antagonist properties.     

In vivo inhibition of dopamine-β-hydroxylase in rat adrenals during exposure to carbon disulphide

Male rats were exposed for a maximum of 4 h to carbon disulphide at atmospheric levels of 1.0—4.0mg/l and the turnover rates of adrenal dopamine was determined by injecting -methyl-p-tyrosine and measuring the rate at which dopamine disappears. Although the level of exposure was significantly higher than the 30.0 g/l permissible limit, or the average occupational exposure, similar or even higher peak exposure values were reported from the viscose rayon industry.After inhibition of tyrosine hydroxylase by -methyl-p-tyrosine, adrenal dopamine contents declined at a slower rate in rats exposed to carbon disulphide than in controls. The reduced rate of dopamine metabolism during exposure to carbon disulphide indicates inhibition of dopamine--hydroxylase in vivo. The size of this effect, which could be detected as soon as 30 min after starting the exposure to carbon disulphide, was dose dependent. The rate of dopamine turnover was still reduced 2 h after the end of a single exposure. However at that time, because of the larger dopamine pool present in the adrenals, the amount of dopamine converted per unit of time was again at pre-exposure levels.S. C. was supported during these studies by grants from the British Wellcome Trust and from the European Medical Research councils.     

The influence of inhibition of catechol-O-methyl transferase or of monoamine oxidase on the extraneuronal metabolism of 3H-(−)-noradrenaline in the rat heart

Summary The extraneuronal metabolism of ³H-(–)-noradrenaline (1 nmol/l) was determined in rat hearts obtained from reserpine-pretreated animals (in the presence of 30 mol/l cocaine).Inhibition of monoamine oxidase (MAO) (by pretreatment of the animals with pargyline) increased the formation of O-methylated metabolites by nearly that amount by which the formation of deaminated metabolites declined; hence, catechol-O-methyl transferase (COMT) seemed to be able to nearly fully compensate for the loss of MAO activity. However, when COMT was inhibited (by the presence of either 1 or 10 mol/l U-O521), the increase in the formation of deaminated metabolites was smaller than the decrease in the formation of O-methylated metabolites; hence, MAO seemed to be unable to fully compensate for the loss of COMT activity.These results are discussed with regard to the hypothesis that the two extraneuronal enzymes co-exist in one compartment. As inhibition of COMT causes a much greater increase in the steady-state tissue/medium ratio for ³H-(–)-noradrenaline than does inhibition of MAO, it is suggested that it is this increase in the intracellular concentration of ³H-(–)-noradrenaline which-by promoting an efflux of the unchanged amine that is proportional to the tissue/medium ratio-actually decreases the net removal of ³H-(–)-noradrenaline from the perfusion fluid.The results are compatible with (but no evidence for) the hypothesis that the two enzymes co-exist in the same extraneuronal compartment.The following abbreviations are used here NMN normetanephrine - DOPEG dihydroxyphenylglycol - DOMA dihydroxymandelic acid - MOPEG methoxyhydroxyphenylglycol - VMA methoxyhydroxymandelic acid - OMDA MOPEG+VMA Supported by the Deutsche Forschungsgemeinschaft     

Safety, tolerability, and pharmacokinetics of etamicastat, a novel dopamine-β-hydroxylase inhibitor, in a rising multiple-dose study in young healthy subjects

Background: Activation of the sympathetic nervous system is an important feature in hypertension and congestive heart failure. A strategy for directly modulating sympathetic nerve function is to reduce the biosynthesis of norepinephrine (noradrenaline) via inhibition of dopamine-β-hydroxylase (DβH).Objective: To assess the safety, tolerability, and pharmacokinetics of etamicastat (BIA 5–453), a new DβH inhibitor, following repeated dosing.Methods: A double-blind, randomized, placebo-controlled study was conducted in healthy young male volunteers. Participants received once-daily doses of placebo or etamicastat 25, 50, 100, 200, 400, or 600 mg, for 10 days.Results: Etamicastat underwent N-acetylation to its metabolite BIA 5–961. Etamicastat and BIA 5–961 maximum concentrations were achieved at 1–3 and 2–4 hours, respectively, after dosing. Elimination half-lives ranged from 18.1 to 25.7 hours for etamicastat and 6.7 to 22.5 hours for BIA 5–961. Both etamicastat and BIA 5-961 followed linear pharmacokinetics. The extent of systemic exposure to etamicastat and BIA 5–961 increased in an approximately dose-proportional manner, and steady-state plasma concentrations were attained up to 9 days of dosing. Etamicastat accumulated in plasma following repeated administration. The mean observed accumulation ratio was 1.3–1.9 for etamicastat and 1.3–1.6 for BIA 5–961. Approximately 40%of the etamicastat dose was recovered in urine in the form of parent compound and BIA 5–961. There was a high variability in pharmacokinetic parameters, attributable to different N-acetyltransferase-2 (NAT2) phenotype. Urinary excretion of norepinephrine decreased following repeated administration of etamicastat. Etamicastat was generally well tolerated. There was no serious adverse event or clinically significant abnormality in clinical laboratory tests, vital signs, or ECG parameters.Conclusion: Etamicastat was well tolerated. Etamicastat undergoes N-acetylation, which is markedly influenced by NAT2 phenotype. NAT2 genotyping could be a step toward personalized medicine for etamicastat.Trial Registration: EudraCT No. 2007-004142-33     

Lack of epinine formation in adrenal medulla and brain of rats during cold exposure and inhibition of dopamine β-hydroxylase

Summary Cold exposure of rats for 4 h and simultaneous inhibition of dopamine -hydroxylase by FLA-63 (25 mg/kg) led to a reduction of the catecholamine content of the adrenal medulla by 46% and of the brain by 68%. Additional injections of 5 mg/kg FLA-63 4 and 9 h after beginning of the experiments, respectively, kept the catecholamine content on this low level (brain) or decreased in further (adrenal medulla). Administration of 5 mg/kg (-)DOPA together with the mono-amine oxidase inhibitor pargyline (50 mg/kg) 24 h after the first injection of FLA-63 stimulated the resynthesis. It amounted for the adrenal medulla to 20 g/kg body weight/8 h and for the brain to 45 ng/g tissue wet weight/8 h. Paper chromatographic analyses of the extracts of adrenal medulla and brain, respectively, performed at each time of the different injections, clearly identified adrenaline, noradrenaline and dopamine (in traces) in the adrenal medulla as well as noradrenaline and dopamine in the brain; epinine on the contrary could not be demonstrated, not even in traces. Since at least 25 ng of epinine can be detected with certainty by our method, it can be concluded that epinine is not formed in amounts greater than 75 ng/pair adrenal glands or 37.5 ng/brain. The present results support the view that the main pathway of adrenaline biosynthesis in the suprarenal medulla and the brain proceeds via noradrenaline and not via epinine.     

Catecholamine modulatory effects of nepicastat (RS-25560-197), a novel,potent and selective inhibitor of dopamine-β-hydroxylase

1. Inhibitory modulation of sympathetic nerve function may have a favourable impact on the progression of congestive heart failure. Nepicastat is a novel inhibitor of dopamine-β-hydroxylase, the enzyme which catalyses the conversion of dopamine to noradrenaline in sympathetic nerves. The in vitro pharmacology and in vivo catecholamine modulatory effects of nepicastat were investigated in the present study.
2. Nepicastat produced concentration-dependent inhibition of bovine (IC₅₀=8.5±0.8 nM) and human (IC₅₀=9.0±0.8  nM)dopamine-β-hydroxylase. The corresponding R-enantiomer (RS-25560-198) was approximately 2–3 fold less potent than nepicastat. Nepicastat had negligible affinity (>10 μM) for twelve other enzymes and thirteen neurotransmitter receptors.
3. Administration of nepicastat to spontaneously hypertensive rats (SHRs) (three consecutive doses of either 3, 10, 30 or 100 mg kg⁻¹, p.o.; 12 h apart) or beagle dogs (0.05, 0.5, 1.5 or 5 mg kg⁻¹, p.o.; b.i.d., for 5 days) produced dose-dependent decreases in noradrenaline content, increases in dopamine content and increases in dopamine/noradrenaline ratio in the artery (mesenteric or renal), left ventricle and cerebral cortex. At the highest dose studied, the decreases in tissue noradrenaline were 47%, 35% and 42% (in SHRs) and 88%, 91% and 96% (in dogs) in the artery, left ventricle and cerebral cortex, respectively. When tested at 30 mg kg⁻¹, p.o., in SHRs, nepicastat produced significantly greater changes in noradrenaline and dopamine content, as compared to the R-enantiomer (RS-25560-198), in the mesenteric artery and left ventricle.
4. Administration of nepicastat (2 mg kg⁻¹, b.i.d, p.o.) to beagle dogs for 15 days produced significant decreases in plasma concentrations of noradrenaline and increases in plasma concentrations of dopamine and dopamine/noradrenaline ratio. The peak reduction (52%) in plasma concentration of noradrenaline and the peak increase (646%) in plasma concentration of dopamine were observed on day-6 and day-7 of dosing, respectively.
5. The findings of this study suggest that nepicastat is a potent, selective and orally active inhibitor of dopamine-β-hydroxylase which produces gradual modulation of the sympathetic nervous system by inhibiting the biosynthesis of noradrenaline. This drug may, therefore, be of value in the treatment of cardiovascular disorders associated with over-activation of the sympathetic nervous system, such as congestive heart failure.

     

The expression and clinical significance of P-GSK3βandβ-catenin in endometrial carcinoma

目的 检测子宫内膜癌组织中P-GSK3β蛋白和β-catenin蛋白表达并探讨其意义.方法 采用微波EliVisionTM免疫组织化学法检测60例子宫内膜癌,32例正常子宫内膜组织中P-GSK3β蛋白和β-catenin蛋白的表达.结果 正常子宫内膜组织中β-catenin蛋白表达正常,子宫内膜癌组织中β-catenin发生异位表达.子宫内膜癌组织中P-GSK3β和异位表达的β-catenin分别定位于细胞浆和细胞核,两者表达明显高于癌旁正常组织(P<0.05).两者的表达与肌层浸润程度、淋巴结转移、临床分期及病理分期有关 (P<0.05);两者之间表达呈正相关(P<0.05).结论 P-GSK3β表达和β-catenin异位表达在子宫内膜癌呈高表达,且与子宫内膜癌的发生、发展和转移密切相关.     

Neurocognitive function in current and ex-users of ecstasy in comparison to both matched polydrug-using controls and drug-naïve controls

Rationale

Despite years of research and much controversy surrounding the effects of ecstasy use, findings are equivocal. Attempting to reduce methodological problems, such as concurrent use of other recreational drugs, could lead to a clearer picture of the effects of ecstasy use on cognitive function.

Objectives

The aim of this study is to investigate the effects of both prolonged abstention from ecstasy use and current use on cognitive function while controlling for the regular use of other recreational drugs used by ecstasy users (alcohol, cannabis, cocaine, amphetamines and lysergic acid diethylamide).

Materials and methods

A range of cognitive functions (including working memory, episodic memory, verbal learning and executive functions) was assessed in 109 participants: 25 current ecstasy users, 28 ex-ecstasy users (abstinent for at least 1 year), 29 polydrug-using controls (matched to both ecstasy-using groups for the use of other recreational drugs) and 27 drug-naïve controls.

Results

There was an overall tendency for impaired verbal learning and memory in both current ecstasy users and polydrug controls. There was also evidence of reduced response inhibition in the current ecstasy users and polydrug controls, which appeared to be related to recency of drug use. However, the majority of tests showed no group differences.

Conclusions

The results suggest that recreational drug use in general, rather than ecstasy use per se, can lead to subtle cognitive impairments and that recent drug use appears to impact strongest on cognitive performance. This study highlights the importance of controlling for the use of all recreational drugs and, in particular, recent drug use when investigating ‘effects of ecstasy’ on cognitive function.

     

Correlation between increased dopamine-β- hydroxylase activity and catecholamine concentration in plasma: Determination of acute changes in sympathetic activity in man

Summary In 11 healthy untrained volunteers the increase in plasma dopamine--hydroxylase (DBH) activity during graded physical exercise has been examined as a true measure of increased activity of the sympathetic nervous system. The correlation between DBH activity, catecholamine concentration (CA) in plasma and heart rate was studied. When work on an electrically braked bicycle ergometer was gradually increased from 12.5 to 100, 200 and 300 watts there was a linear increase in DBH activity and heart rate; the increase in CA concentrations followed an exponential function. The peak values for DBH activity and CA concentration in plasma after the 300 watt work load (as percentages of the resting levels) were 130±3% and 820±71%, respectively; the adrenaline concentration in plasma increased only to 150±19% (p>0.05). There were significant correlations between heart rate and work load, DBH and work load and log CA and work load. The data imply direct correlations between heart rate and DBH, heart rate and log CA and DBH and log CA. The exponential increase in noradrenaline concentration in plasma might be due either to a greater net overflow from sympathetic nerve endings, and/or to increased secretion by the adrenal medulla. In the latter case, the release of noradrenaline would not be accompanied by secretion either of adrenaline or DBH. After work ceased there were sharp falls in heart rate and CA concentration, which indicate an immediate drop in sympathetic activity. DBH activity in plasma returned to normal very slowly; it reached half maximum values after 20 – 22 min. It is concluded that increased sympathetic activity in man can be estimated in vivo as changes in DBH and/or CA concentration in plasma. In contrast, a rapid decrease in sympathetic activity is directly reflected only by a rapid fall in the plasma concentrations of CA.Part of this work was presented at the Symposion on Hypertension. Mainz, November 1973; Wietholdet al., 1973.Supported by a grant from the Bundesinstitut für Sportwissenschaft, Köln.     

Antiproliferative and teratogenic activities of the bishemisuccinates of 7α-hydroxycholesterol and 7β-hydroxycholesterol

1. The bishemisuccinates of 7α-hydroxycholesterol and 7β-hydroxycholesterol induced a dose-dependent decrease in the incorporation of [³H]thymidine into fibroblast cell line (3T3), macrophagelike cell line (P388D1) and rat hepatoma cell line (H35). Concomitantly there was a reduction in cell viability.2. The 3T3 cells were less susceptible than P388D1 and H35 cells.3. The aforementioned derivatives of 7α-hydroxycholesterol and 7β-hydroxycholesterol produced malformations in cultured embryos including abnormalities in yolk sac circulation, heartbeat, body axis, otic vesicle, branchial apparatus, cranial neural tube and forelimb bud. Axial length and somite number were reduced.     

The prospects of brain — computer interface applications in children

The restoring of motor functions in adults through brain-computer interface applications is widely studied in the contemporary literature. But there is a lack of similar analyses and research on the application of brain-computer interfaces in the neurorehabilitation of children. There is a need for expanded knowledge in the aforementioned area. This article aims at investigating the extent to which the available opportunities in the area of neurorehabilitation and neurological physiotherapy of children with severe neurological deficits using brain-computer interfaces are being applied, including our own concepts, research and observations.     

The Pharmacokinetics of β-Cyclodextrin and Hydroxypropyl-β-cyclodextrin in the Rat

Hydroxypropyl--cyclodextrin was analyzed by HPLC using postcolumn complexation with phenolphthalein and negative colorimetric detection, with a detection limit of 20 µg/ml. The pharmacokinetics of -cyclodextrin and of hydroxypropyl--cyclodextrin were studied after intravenous administration to permanently cannulated rats. The pharmacokinetic behavior of both cyclodextrins was similar to that of inulin, showing rapid distribution over extracellular fluids. Elimination occurred through glomerular filtration. When a dose of 200 mg/kg -cyclodextrin was administered the elimination rate was decreased, probably as a result of nephrotoxicity of -cyclodextrin. Within 24 hr after administration most of the cyclodextrin dose was recovered unchanged in urine. After oral administration, only insignificant amounts of intact -cyclodextrin were absorbed from the gastrointestinal tract.     

The role of pro-/anti-inflammation imbalance in Aβ42 accumulation of rat brain co-exposed to fine particle matter and sulfur dioxide

Taiyuan is a center of coal-based electricity production and many chemicals industries, where mixtures of sulfur dioxide (SO₂) and particulate matter may be more prominent. The focus of the present study was to determine if there is a link between adverse effects in the brain and the combined-exposure to SO₂ and fine particulate matter (PM_2.5). Rats were exposed alternately to PM_2.5 with different dosages (1.5, 6.0 and 24.0?mg/kg body weight) and SO₂ at the level of 5.6?mg/m³. The results showed that the combined exposure to PM_2.5 and SO₂ enhanced the mRNA expression and protein level of TNF-α and IL-6 in rat cortex and hippocampus relative to the control, SO₂ and PM_2.5 alone. Instead, TGF-β1 mRNA and protein level were down-regulated in the brain. Additionally, PM_2.5 at medium and/or high dose caused marked increase in Aβ42 level and PM_2.5?+?SO₂ induced further increase of Aβ42 level in the cortex and hippocampus. It suggests that SO₂ and PM_2.5 can synergistically exert inflammation responses and induce Aβ42 accumulation in the brain. Also, it is notable that the Aβ42 accumulation of rat cortex and hippocampus were closely associated with pro-/anti-inflammatory cytokines ratio. These results clearly demonstrated that the combined exposure to PM_2.5 and SO₂ can induce the imbalance of pro-/anti-inflammatory cytokine, resulting in Aβ42 accumulation of rat brain cortex and hippocampus.     

Expression of cysteinyl leukotriene receptors in human traumatic brain injury and brain tumors

Cysteinyl leukotrienes (CysLTs) are potent proinflammatory mediators. Till now only CysLT receptor 1 (CysLT1) and CysLT receptor 2 (CysLT2) have been cloned. Although the existence of CysLT1 and CysLT2 in the brain has been demonstrated by Northern blot and RT-PCR analyses, the exact location of the receptors in the brain remains unknown. The objective