Delayed O-methylation of L-DOPA in MB-COMT-deficient mice after oral administration of L-DOPA and carbidopa

1.Catechol-O-methyltransferase (COMT) is involved in the O-methylation of l-DOPA, dopamine, and other catechols. The enzyme is expressed in two isoforms: soluble (S-COMT), which resides in the cytoplasm, and membrane-bound (MB-COMT), which is anchored to intracellular membranes. 2.To obtain specific information on the functions of COMT isoforms, we studied how a complete MB-COMT deficiency affects the total COMT activity in the body, peripheral l-DOPA levels, and metabolism after l-DOPA (10mg kg(-1)) plus carbidopa (30mg kg(-1)) administration by gastric tube in wild-type (WT) and MB-COMT-deficient mice. l-DOPA and 3-O-methyl-l-DOPA (3-OMD) levels were assayed in plasma, duodenum, and liver. 3.We showed that the selective lack of MB-COMT did not alter the total COMT activity, COMT enzyme kinetics, l-DOPA levels, or the total O-methylation of l-DOPA but delayed production of 3-OMD in plasma and peripheral tissues.Peer reviewe

Catechol-O-methyltransferase (COMT) protein expression and activity after dopaminergic and noradrenergic lesions of the rat brain

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Reduced natriuretic response to acute sodium loading in COMT Gene deleted mice

BACKGROUND: The intrarenal natriuretic hormone dopamine (DA) is metabolised by catechol-O-methyltransferase (COMT) and monoamine oxidase (MAO). Inhibition of COMT, as opposed to MAO, results in a potent natriuretic response in the rat. The present study in anaesthetized homozygous and heterozygous COMT gene deleted mice attempted to further elucidate the importance of COMT in renal DA and sodium handling. After acute intravenous isotonic sodium loading, renal function was followed. RESULTS: COMT activity in heterozygous mice was about half of that in wild type mice and was zero in the homozygous mice. MAO activity did not differ between the genotypes. Urinary sodium excretion increased 10-fold after sodium loading in wild type mice. In heterozygous and homozygous mice, the natriuretic effects of sodium loading were only 29 % and 39 %, respectively, of that in wild type mice. Arterial pressure and glomerular filtration rate did not differ between genotypes. Baseline norepinephrine and DA excretions in urine were elevated in the homozygous, but not in heterozygous, COMT gene deleted mice. Urinary DA excretion increased after isotonic sodium loading in the wild type mice but not in the COMT gene deleted mice. CONCLUSIONS: Mice with reduced or absent COMT activity have altered metabolism of catecholamines and are unable to increase renal DA activity and produce normal natriuresis in response to acute sodium loading. The results support the hypothesis that COMT has an important role in the DA-mediated regulation of renal sodium excretion

Transcriptional profiling of C57 and DBA strains of mice in the absence and presence of morphine

BACKGROUND: The mouse C57BL/6 (C57) and DBA/2J (DBA) inbred strains differ substantially in many aspects of their response to drugs of abuse. The development of microarray analyses represents a genome-wide method for measuring differences across strains, focusing on expression differences. In the current study, we carried out microarray analysis in C57 and DBA mice in the nucleus accumbens of drug-naïve and morphine-treated animals. RESULTS: We identified mRNAs with altered expression between the two strains. We validated the mRNA expression changes of several such mRNAs, including Gnb1, which has been observed to be regulated by several drugs of abuse. In addition, we validated alterations in the enzyme activity of one mRNA product, catechol-O-methyltransferase (Comt). Data mining of expression and behavioral data indicates that both Gnb1 and Comt expression correlate with aspects of drug response in C57/DBA recombinant inbred strains. Pathway analysis was carried out to identify pathways showing significant alterations as a result of treatment and/or due to strain differences. These analyses identified axon guidance genes, particularly the semaphorins, as showing altered expression in the presence of morphine, and plasticity genes as showing altered expression across strains. Pathway analysis of genes showing strain by treatment interaction suggest that the phosphatidylinositol signaling pathway may represent an important difference between the strains as related to morphine exposure. CONCLUSION: mRNAs with differing expression between the two strains could potentially contribute to strain-specific responses to drugs of abuse. One such mRNA is Comt and we hypothesize that altered expression of Comt may represent a potential mechanism for regulating the effect of, and response to, multiple substances of abuse. Similarly, a role for Gnb1 in responses to multiple drugs of abuse is supported by expression data from our study and from other studies. Finally, the data support a role for semaphorin signaling in morphine effects, and indicate that altered expression of genes involved in phosphatidylinositol signaling and plasticity might also affect the altered drug responses in the two strains

Pharmacokinetics and Pharmacodynamics of Entacapone and Tolcapone after Acute and Repeated Administration: A Comparative Study in the Rat

ABSTRACT Two catechol-O-methyltransferase (COMT) inhibitors, entacapone and tolcapone, were compared in the rat to elucidate the actual differences between their pharmacokinetics and pharmacodynamics after single and repeated administration. Their inhibitory potencies were also compared in vitro. After intravenous administration (3 mg/kg), the elimination half-life (t 1/2␤ ) of entacapone (0.8 h) was clearly shorter than that of tolcapone (2.9 h). The striatum/serum ratio of tolcapone was 3-fold higher than that of entacapone. After a single oral dose (10 mg/kg), both entacapone and tolcapone produced an equal maximal degree of COMT inhibition in peripheral tissues, but tolcapone inhibited striatal COMT more effectively than did entacapone. After the 7-day treatment (10 mg/kg twice daily), COMT activity had recovered to a level of 67 to 101% of control within 8 h after the last dose of entacapone. In tolcapone-treated animals, there was still extensive COMT inhibition present in peripheral tissues, and the degree of inhibition was higher than that attained after a single dose. The pharmacokinetic-pharmacodynamic modeling revealed that a plateau of COMT inhibition near the maximal attainable inhibition was reached already by plasma concentrations below 2000 ng/ml, both with entacapone and tolcapone. Entacapone and tolcapone inhibited equally rat liver COMT in vitro with K i values of 10.7 and 10.0 nM, respectively. In conclusion, tolcapone has a longer duration of action and a better brain penetration than entacapone. The results also suggest that peripheral COMT is inhibited continuously when tolcapone is dosed at 12-h intervals, but this was not seen with entacapone. The second-generation catechol-O-methyltransferase (COMT, EC 2.1.1.6) inhibitors, entacapone and tolcapone, are indicated as adjuncts to standard levodopa-dopa decarboxylase inhibitor therapy in Parkinson's disease. They increase the bioavailability of levodopa by inhibiting its peripheral metabolism to an inactive metabolite, 3-O-methyldopa. COMT inhibitors improve the efficacy of the levodopa-dopa decarboxylase inhibitor therapy by prolonging the duration of action and the clinical benefit of levodopa Entacapone and tolcapone apparently behave differently both in experimental animals and humans. However, as a rule, entacapone and tolcapone have been studied only individually; their pharmacokinetics and pharmacodynamics have not been compared thoroughly after single and repeated dosing. Actually, very little is known about their pharmacodynamics in different tissues after repeated dosing. Furthermore, only the relationship between the plasma drug concentration and COMT activity in erythrocytes has been studied previously A few available studies on entacapone and tolcapone in rats suggest that entacapone is eliminated faster than tolcapone and its oral bioavailability is lower than that of tolcapone. After intravenous administration of 10 mg/kg tolcapone, the t 1/2 was 0.9 h and total clearance 470 ml ϫ h Ϫ1 ϫ kg Ϫ1 . The oral bioavailability was 48% for 20 mg/kg and 56% for 40 mg/kg The time course of COMT activity in different tissues has ABBREVIATIONS. COMT, catechol-O-methyltransferase; t 1/2␤ , elimination half-life (␤-phase); S-COMT; soluble catechol-O-methyltransferase; MB-COMT, membrane-bound catechol-O-methyltransferase; AUE, area under the effect-time curve; AUC, area under the plasma drug concentration-time curve; C 0 , initial plasma concentration; E 0 , baseline effect; E max , maximum attainable effect

Combination of CDNF and Deep Brain Stimulation Decreases Neurological Deficits in Late-stage Model Parkinson's Disease

Several neurotrophic factors ( NTF) are shown to be neuroprotective and neurorestorative in pre-clinical animal models for Parkinson's disease ( PD), particularly in models where striatal dopamine neuron innervation partially exists. The results of clinical trials on late-stage patients have been modest. Subthalamic deep brain stimulation ( STN DBS) is a proven treatment for a selected group of advanced PD patients. The cerebral dopamine neurotrophic factor ( CDNF) is a promising therapeutic protein, but its effects in animal models of late-stage PD have remained under-researched. The interactions of NTF and STN DBS treatments have not been studied before. We found that a nigral CDNF protein alone had only a marginal effect on the behavioral deficits in a late-stage hemiparkinsonian rat model ( 6-OHDA MFB). However, CDNF improved the effect of acute STN DBS on front limb use asymmetry at 2 and 3 weeks after CDNF injection. STN lesion-modeling chronic stimulation-had an additive effect in reducing front limb use in the cylinder test and apomorphine-induced rotation. The combination of CDNF and acute STN DBS had a favorable effect on striatal tyrosine hydroxylase. This study presents a novel additive beneficial effect of NTF and STN DBS, which might be explained by the interaction of DBS-induced endogenous NTFs and exogenously injected CDNF. SNpc can be reached via similar trajectories used in clinical STN DBS, and this interaction is an important area for future studies. (C) 2018 The Authors. Published by Elsevier Ltd on behalf of IBRO. This is an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/).Peer reviewe

Epistasis between polymorphisms in COMT, ESR1, and GCH1 influences COMT enzyme activity and pain

Abnormalities in the enzymatic activity of catechol-O-methyltransferase (COMT) contribute to chronic pain conditions, such as temporomandibular disorders (TMD). Thus, we sought to determine the effects of polymorphisms in COMT and functionally-related pain genes in the COMT pathway (estrogen receptor 1: ESR1, guanosine-5-triphosphate cyclohydrolase 1: GCH1, methylenetetrahydrofolate reductase: MTHFR) on COMT enzymatic activity, musculoskeletal pain, and pain-related intermediate phenotypes among TMD cases and healthy controls. Results demonstrate that the COMT rs4680 (val158met) polymorphism is most strongly associated with outcome measures, such that individuals with the minor A allele (met) exhibit reduced COMT activity, increased TMD risk, and increased musculoskeletal pain. Epistatic interactions were observed between the COMT rs4680 polymorphism and polymorphisms in GCH1 and ESR1. Among individuals with the COMT met allele, those with two copies of the GCH1 rs10483639 minor G allele exhibit normalized COMT activity and increased mechanical pain thresholds. Among individuals with the COMT val allele, those with two copies of the ESR1 rs3020377 minor A allele exhibit reduced COMT activity, increased bodily pain, and poorer self-reported health. These data reveal that the GCH1 minor G allele confers a protective advantage among met carriers, while the ESR1 minor A allele is disadvantageous among val carriers. Furthermore, these data suggest that the ability to predict the downstream effects of genetic variation on COMT activity is critically important to understanding the molecular basis of chronic pain conditions

Subacute administration of both methcathinone and manganese causes basal ganglia damage in mice resembling that in methcathinone abusers

An irreversible extrapyramidal syndrome occurs in man after intravenous abuse of "homemade" methcathinone (ephedrone, Mcat) that is contaminated with manganese (Mn) and is accompanied by altered basal ganglia function. Both Mcat and Mn can cause alterations in nigrostriatal function but it remains unknown whether the effects of the 'homemade' drug seen in man are due to Mcat or to Mn or to a combination of both. To determine how toxicity occurs, we have investigated the effects of 4-week intraperitoneal administration of Mn (30 mg/kg t.i.d) and Mcat (100 mg/kg t.i.d.) given alone, on the nigrostriatal function in male C57BL6 mice. The effects were compared to those of the 'homemade' mixture which contained about 7 mg/kg of Mn and 100 mg/kg of Mcat. Motor function, nigral dopaminergic cell number and markers of pre- and postsynaptic dopaminergic neuronal integrity including SPECT analysis were assessed. All three treatments had similar effects on motor behavior and neuronal markers. All decreased motor activity and induced tyrosine hydroxylase positive cell loss in the substantia nigra. All reduced I-123-epidepride binding to D2 receptors in the striatum. Vesicular monoamine transporter 2 (VMAT2) binding was not altered by any drug treatment. However, Mcat treatment alone decreased levels of the dopamine transporter (DAT) and Mn alone reduced GAD immunoreactivity in the striatum. These data suggest that both Mcat and Mn alone could contribute to the neuronal damage caused by the 'homemade' mixture but that both produce additional changes that contribute to the extrapyramidal syndrome seen in man.Peer reviewe

Gene therapy with AAV2-CDNF provides functional benefits in a rat model of Parkinson's disease

Cerebral dopamine neurotrophic factor (CDNF) protein has been shown to protect the nigrostriatal dopaminergic pathway when given as intrastriatal infusions in rat and mouse models of Parkinson's disease (PD). In this study, we assessed the neuroprotective effect of CDNF delivered with a recombinant adeno-associated viral (AAV) serotype 2 vector in a rat 6-hydroxydopamine (6-OHDA) model of PD. AAV2 vectors encoding CDNF, glial cell line-derived neurotrophic factor (GDNF), or green fluorescent protein were injected into the rat striatum. Protein expression analysis showed that our AAV2 vector efficiently delivered the neurotrophic factor genes into the brain and gave rise to a long-lasting expression of the proteins. Two weeks after AAV2 vector injection, 6-OHDA was injected into the rat striatum, creating a progressive degeneration of the nigrostriatal dopaminergic system. Treatment with AAV2-CDNF resulted in a marked decrease in amphetamine-induced ipsilateral rotations while it provided only partial protection of tyrosine hydroxylase (TH)-immunoreactive cells in the rat substantia nigra pars compacta and TH-reactive fibers in the striatum. Results from this study provide additional evidence that CDNF can be considered a potential treatment of Parkinson's disease.Peer reviewe

Application of non-HDL cholesterol for population-based cardiovascular risk stratification: results from the Multinational Cardiovascular Risk Consortium.

BACKGROUND: The relevance of blood lipid concentrations to long-term incidence of cardiovascular disease and the relevance of lipid-lowering therapy for cardiovascular disease outcomes is unclear. We investigated the cardiovascular disease risk associated with the full spectrum of bloodstream non-HDL cholesterol concentrations. We also created an easy-to-use tool to estimate the long-term probabilities for a cardiovascular disease event associated with non-HDL cholesterol and modelled its risk reduction by lipid-lowering treatment. METHODS: In this risk-evaluation and risk-modelling study, we used Multinational Cardiovascular Risk Consortium data from 19 countries across Europe, Australia, and North America. Individuals without prevalent cardiovascular disease at baseline and with robust available data on cardiovascular disease outcomes were included. The primary composite endpoint of atherosclerotic cardiovascular disease was defined as the occurrence of the coronary heart disease event or ischaemic stroke. Sex-specific multivariable analyses were computed using non-HDL cholesterol categories according to the European guideline thresholds, adjusted for age, sex, cohort, and classical modifiable cardiovascular risk factors. In a derivation and validation design, we created a tool to estimate the probabilities of a cardiovascular disease event by the age of 75 years, dependent on age, sex, and risk factors, and the associated modelled risk reduction, assuming a 50% reduction of non-HDL cholesterol. FINDINGS: Of the 524 444 individuals in the 44 cohorts in the Consortium database, we identified 398 846 individuals belonging to 38 cohorts (184 055 [48·7%] women; median age 51·0 years [IQR 40·7-59·7]). 199 415 individuals were included in the derivation cohort (91 786 [48·4%] women) and 199 431 (92 269 [49·1%] women) in the validation cohort. During a maximum follow-up of 43·6 years (median 13·5 years, IQR 7·0-20·1), 54 542 cardiovascular endpoints occurred. Incidence curve analyses showed progressively higher 30-year cardiovascular disease event-rates for increasing non-HDL cholesterol categories (from 7·7% for non-HDL cholesterol <2·6 mmol/L to 33·7% for ≥5·7 mmol/L in women and from 12·8% to 43·6% in men; p<0·0001). Multivariable adjusted Cox models with non-HDL cholesterol lower than 2·6 mmol/L as reference showed an increase in the association between non-HDL cholesterol concentration and cardiovascular disease for both sexes (from hazard ratio 1·1, 95% CI 1·0-1·3 for non-HDL cholesterol 2·6 to <3·7 mmol/L to 1·9, 1·6-2·2 for ≥5·7 mmol/L in women and from 1·1, 1·0-1·3 to 2·3, 2·0-2·5 in men). The derived tool allowed the estimation of cardiovascular disease event probabilities specific for non-HDL cholesterol with high comparability between the derivation and validation cohorts as reflected by smooth calibration curves analyses and a root mean square error lower than 1% for the estimated probabilities of cardiovascular disease. A 50% reduction of non-HDL cholesterol concentrations was associated with reduced risk of a cardiovascular disease event by the age of 75 years, and this risk reduction was greater the earlier cholesterol concentrations were reduced. INTERPRETATION: Non-HDL cholesterol concentrations in blood are strongly associated with long-term risk of atherosclerotic cardiovascular disease. We provide a simple tool for individual long-term risk assessment and the potential benefit of early lipid-lowering intervention. These data could be useful for physician-patient communication about primary prevention strategies. FUNDING: EU Framework Programme, UK Medical Research Council, and German Centre for Cardiovascular Research