Publications by authors named "Kamila Syslova"

30 Publications

  • Page 1 of 1

Alterations in Rat Accumbens Dopamine, Endocannabinoids and GABA Content During WIN55,212-2 Treatment: The Role of Ghrelin.

Int J Mol Sci 2020 Dec 28;22(1). Epub 2020 Dec 28.

Department of Pharmacology, Third Faculty of Medicine, Charles University, Ruska 87, 100 34 Prague 10, Czech Republic.

The endocannabinoid/CB1R system as well as the central ghrelin signalling with its growth hormone secretagogoue receptors (GHS-R1A) are importantly involved in food intake and reward/reinforcement processing and show distinct overlaps in distribution within the relevant brain regions including the hypothalamus (food intake), the ventral tegmental area (VTA) and the nucleus accumbens (NAC) (reward/reinforcement). The significant mutual interaction between these systems in food intake has been documented; however, the possible role of ghrelin/GHS-R1A in the cannabinoid reinforcement effects and addiction remain unclear. Therefore, the principal aim of the present study was to investigate whether pretreatment with GHS-R1A antagonist/JMV2959 could reduce the CB1R agonist/WIN55,212-2-induced dopamine efflux in the nucleus accumbens shell (NACSh), which is considered a crucial trigger impulse of the addiction process. The synthetic aminoalklylindol cannabinoid WIN55,212-2 administration into the posterior VTA induced significant accumbens dopamine release, which was significantly reduced by the 3 mg/kg i.p. JMV2959 pretreatment. Simultaneously, the cannabinoid-increased accumbens dopamine metabolic turnover was significantly augmented by the JMV2959 pretreament. The intracerebral WIN55,212-2 administration also increased the endocannabinoid arachidonoylethanolamide/anandamide and the 2-arachidonoylglycerol/2-AG extracellular levels in the NACSh, which was moderately but significantly attenuated by the JMV2959 pretreatment. Moreover, the cannabinoid-induced decrease in accumbens γ-aminobutyric acid/gamma-aminobutyric acid levels was reversed by the JMV2959 pretreatment. The behavioural study in the LABORAS cage showed that 3 mg/kg JMV2959 pretreatment also significantly reduced the systemic WIN55,212-2-induced behavioural stimulation. Our results demonstrate that the ghrelin/GHS-R1A system significantly participates in the rewarding/reinforcing effects of the cannabinoid/CB1 agonist that are involved in cannabinoid addiction processing.
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http://dx.doi.org/10.3390/ijms22010210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7795825PMC
December 2020

Ghrelin receptor antagonism of fentanyl-induced conditioned place preference, intravenous self-administration, and dopamine release in the nucleus accumbens in rats.

Addict Biol 2020 11 6;25(6):e12845. Epub 2019 Nov 6.

Department of Pharmacology, Third Faculty of Medicine, Charles University, Czech Republic.

The extended occurrence of fentanils abuse associated with the dramatic increase in opioid fatal overdoses and dependence strongly emphasizes insufficiencies in opioid addiction treatment. Recently, the growth hormone secretagogue receptor (GHS-R1A) antagonism was proposed as a promising mechanism for drug addiction therapy. However, the role of GHS-R1A and its endogenous ligand ghrelin in opioid abuse is still unclear. Therefore, the aim of our study was to clarify whether the GHS-R1A antagonist JMV2959 could reduce the fentanyl-induced conditioned place preference (CPP), the fentanyl intravenous self-administration (IVSA), and the tendency to relapse, but also whether JMV2959 could significantly influence the fentanyl-induced dopamine efflux in the nucleus accumbens (NAC) in rats, that importantly participates in opioids' reinforcing effects. Following an ongoing fentanyl self-administration, JMV2959 3 mg/kg was administered intraperitoneally 20 minutes before three consequent daily 360-minute IVSA sessions under a fixed ratio FR1, which significantly reduced the number of active lever-pressing, the number of infusions, and the fentanyl intake. Pretreatment with JMV2959 also reduced the fentanyl-seeking/relapse-like behaviour tested in rats on the 12 day of the forced abstinence period. Pretreatment with JMV2959 significantly and dose-dependently reduced the manifestation of fentanyl-CPP. The fentanyl-CPP development was reduced after the simultaneous administration of JMV2959 with fentanyl during conditioning. The JMV2959 significantly reduced the accumbens dopamine release induced by subcutaneous and intravenous fentanyl. Simultaneously, it affected the concentration of byproducts associated with dopamine metabolism in the NAC. Our findings suggest that GHS-R1A importantly participates in the rewarding/reinforcing effects of fentanyl.
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http://dx.doi.org/10.1111/adb.12845DOI Listing
November 2020

Blood-based molecular signature of Alzheimer's disease via spectroscopy and metabolomics.

Clin Biochem 2019 Oct 4;72:58-63. Epub 2019 Apr 4.

Department of Analytical Chemistry, University of Chemistry and Technology Prague, Technická 5, 166 28 Prague 6, Czech Republic.

Objectives: With over 35 million cases worldwide, Alzheimer's disease (AD) represents the main cause of dementia. The differentiation of AD from other types of dementia is challenging and its early diagnosis is complicated. The established biomarkers are not only based on the invasive collection of cerebrospinal fluid, but also lack sufficient sensitivity and specificity. Therefore, much current effort is aimed at the identification of new biomarkers of AD in peripheral blood.

Design And Methods: We focused on blood-based analyses using chiroptical spectroscopy (Raman optical activity, electronic circular dichroism) supplemented with conventional vibrational spectroscopy (infrared, Raman) and metabolomics (high-performance liquid chromatography with a high-resolution mass detection).

Results: This unique approach enabled us to identify the spectral pattern of AD and variations in metabolite levels. Subsequent linear discriminant analysis of the spectral data resulted in differentiation between the AD patients and control subjects.

Conclusions: It may be stated that this less invasive approach has strong potential for the identification of disease-related changes within essential plasmatic biomolecules and metabolites.
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http://dx.doi.org/10.1016/j.clinbiochem.2019.04.004DOI Listing
October 2019

Monitoring of kynurenine pathway metabolites, neurotransmitters and their metabolites in blood plasma and brain tissue of individuals with latent toxoplasmosis.

J Pharm Biomed Anal 2019 Jun 18;170:139-152. Epub 2019 Mar 18.

National Institute of Mental Health, Topolová 748, 250 67, Klecany, Czech Republic.

The aim of the presented work was to develop a highly sensitive, accurate and rapid analytical method for the determination of concentration levels of tryptophan and its metabolites of kynurenine catabolic pathway, as well as neurotransmitters and their metabolites in complex biological matrices (brain tissue and blood plasma). The developed analytical method consists of analytes separation from the biological matrices by protein precipitation (blood plasma) or solvent extraction (brain tissue), derivatization of the analytes and their detection by high-performance liquid chromatography combined with mass spectrometry. Individual steps of the whole process were optimized and the method was validated in the terms of selectivity, linearity (R≥0.980), precision (RSD ≤ 13.3%), recovery (≥82.0%), limit of detection (1.8 ng/mL of blood plasma, 2.2 pg/mg of brain tissue) and limit of quantification (2.5 ng/mL of blood plasma, 2.8 pg/mg of brain tissue). The method was subsequently verified by an animal study, where the concentration levels of the analytes in biological matrices (blood plasma and brain tissue) of T. gondii - infected rats and control animals were compared. All the data obtained from the animal study were statistically evaluated. Increased concentration levels of kynurenine catabolic pathway metabolites (e.g. kynurenine, 3-hydroxykynurenine, quinolinic acid) were observed in the case of T. gondii - infected rats in contrast to the control group. The opposite effect was determined in the case of serotonin and its metabolite 5-hydroxyindoleacetic acid, where higher concentration levels were found in blood plasma of healthy subjects. Finally, Principal Component Analysis (PCA) was utilized for a score plot formation. PCA score plots have demonstrated the similarities of individuals within each group and the differences among the groups.
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http://dx.doi.org/10.1016/j.jpba.2019.03.039DOI Listing
June 2019

Chiroptical spectroscopy and metabolomics for blood-based sensing of pancreatic cancer.

Chirality 2018 05 23;30(5):581-591. Epub 2018 Feb 23.

Department of Analytical Chemistry, University of Chemistry and Technology Prague, Prague 6, Czech Republic.

To enable the early diagnosis of pancreatic cancer, the search for and definition of reliable biomarkers remain a subject of great interest, with the specificity and sensitivity of the currently used biomarkers being below the required values. We tested a novel diagnostic approach for pancreatic cancer based on the specific molecular signature of blood plasma components. To acquire more detailed structural information, structure-sensitive chiroptical methods (electronic circular dichroism and Raman optical activity) were supplemented by conventional Raman and infrared spectroscopies. The obtained spectra were subsequently processed by linear discriminant analysis yielding high values of specificity and sensitivity. In addition, to monitor not only large biomolecules as potential biomarkers but also those of low molecular weight, we conducted an analysis of blood plasma samples by using metabolomics. The achieved results suggest a panel of promising biomarkers for a reliable detection of pancreatic cancer.
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http://dx.doi.org/10.1002/chir.22834DOI Listing
May 2018

Alterations in Rat Accumbens Endocannabinoid and GABA Content during Fentanyl Treatment: The Role of Ghrelin.

Int J Mol Sci 2017 11 22;18(11). Epub 2017 Nov 22.

Laboratory of Medicinal Diagnostics, Department of Organic Technology ICT, Technicka 5, 166 28 Prague 6, Czech Republic.

The opioid-induced rise of extracellular dopamine, endocannabinoid anandamide and γ-aminobutyric acid (GABA) concentrations triggered by opioids in the nucleus accumbens shell (NACSh) most likely participate in opioid reward. We have previously demonstrated that systemic administration of ghrelin antagonist (JMV2959) significantly decreased morphine-induced dopamine and anandamide (-arachidonoylethanolamine, AEA) increase in the NACSh. Fentanyl is considered as a µ-receptor-selective agonist. The aim of this study was to test whether JMV2959, a growth hormone secretagogue receptor (GHS-R1A) antagonist, can influence the fentanyl-induced effects on anandamide, 2-arachidonoylglycerol (2-AG) and GABA in the NACSh and specify the involvement of GHS-R1A located in the ventral tegmental area (VTA) and nucleus accumbens (NAC). Using in vivo microdialysis in rats, we have found that pre-treatment with JMV2959 reversed dose dependently fentanyl-induced anandamide increases in the NACSh, resulting in a significant AEA decrease and intensified fentanyl-induced decreases in accumbens 2-AG levels, with both JMV2959 effects more expressed when administered into the NACSh in comparison to the VTA. JMV2959 pre-treatment significantly decreased the fentanyl-evoked accumbens GABA efflux and reduced concurrently monitored fentanyl-induced behavioural stimulation. Our current data encourage further investigation to assess if substances affecting GABA or endocannabinoid concentrations and action, such as GHS-R1A antagonists, can be used to prevent opioid-seeking behaviour.
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http://dx.doi.org/10.3390/ijms18112486DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5713452PMC
November 2017

Trans-generational neurochemical modulation of methamphetamine in the adult brain of the Wistar rat.

Arch Toxicol 2017 Oct 5;91(10):3373-3384. Epub 2017 May 5.

Department of Normal, Pathological and Clinical Physiology, Third Faculty of Medicine, Charles University, Prague, Czech Republic.

Chronic methamphetamine (METH) abuse has been shown to elicit strong neurotoxic effects. Yet, with an increasing number of children born to METH abusing mothers maturing into adulthood, one important question is how far do the neurotoxic effects of METH alter various neurotransmitter systems in the adult METH-exposed offspring. The purpose of this study was to investigate long-term trans-generational neurochemical changes, following prenatal METH exposure, in the adult Wistar rat brain. METH or saline (SAL-control animals) was administered to pregnant dams throughout the entire gestation period (G0-G22). At postnatal day 90, dopamine, serotonin, glutamate and GABA were measured in the adult brain before (baseline) and after a METH re-administration using in vivo microdialysis and liquid chromatography/mass spectrometry. The results show that METH-exposure increased basal levels of monoamines and glutamate, but decreased GABA levels in all measured brain regions. Acute challenge with METH injection in the METH-exposed group induced a lower increase in the monoamine system relative to the increase in the GABAergic and glutamatergic system. The data show that prenatal METH exposure has strong effects on the monoaminergic, GABAergic and glutamatergic system even when exposure to METH was limited to the prenatal phase. Toxicological effects of METH have therefore longer lasting effects as currently considered and seem to affect the excitatory-inhibitory balance in the brain having strong implications for cognitive and behavioral functioning.
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http://dx.doi.org/10.1007/s00204-017-1969-yDOI Listing
October 2017

Distributions of therapeutically promising neurosteroids in cellular membranes.

Chem Phys Lipids 2017 03 30;203:78-86. Epub 2016 Dec 30.

J. Heyrovský Institute of Physical Chemistry, Academy of Sciences of the Czech Republic, Dolejškova 3, 182 23 Prague, Czech Republic. Electronic address:

Interactions of two neurosteroids, inhibiting membrane-bound N-Methyl-d-aspartate receptors, with phospholipid membranes are studied. Namely, endogenous pregnanolone sulfate is compared with pregnanolone glutamate, the latter being a novel synthetic steroidal inhibitor of these receptors with potential pharmaceutical use. Molecular-level details of steroid-phospholipid membranes interactions are scrutinized employing molecular dynamics simulations supported by quantum chemical calculations to assess steroid lipophilicity. Moreover, permeability of both species across membranes is experimentally evaluated by immobilized artificial membrane chromatography. We demonstrate that while there is no significant difference of lipophilicity and membrane permeability between the two steroids, they differ significantly regarding detailed localization in phospholipid membranes. The bulky glutamate moiety of pregnanolone glutamate is flexible and well exposed to the water phase while the sulfate group of pregnanolone sulfate is hidden in the membrane headgroup region. This dissimilarity of behavior in membranes can potentially account for the observed different activities of the two steroids toward membrane-bound N-Methyl-d-aspartate receptors.
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http://dx.doi.org/10.1016/j.chemphyslip.2016.12.004DOI Listing
March 2017

Repeated intraspecific divergence in life span and aging of African annual fishes along an aridity gradient.

Evolution 2017 Feb 7;71(2):386-402. Epub 2016 Dec 7.

Institute of Vertebrate Biology, Academy of Sciences of the Czech Republic, Květná 8, 603 65, Brno, Czech Republic.

Life span and aging are substantially modified by natural selection. Across species, higher extrinsic (environmentally related) mortality (and hence shorter life expectancy) selects for the evolution of more rapid aging. However, among populations within species, high extrinsic mortality can lead to extended life span and slower aging as a consequence of condition-dependent survival. Using within-species contrasts of eight natural populations of Nothobranchius fishes in common garden experiments, we demonstrate that populations originating from dry regions (with short life expectancy) had shorter intrinsic life spans and a greater increase in mortality with age, more pronounced cellular and physiological deterioration (oxidative damage, tumor load), and a faster decline in fertility than populations from wetter regions. This parallel intraspecific divergence in life span and aging was not associated with divergence in early life history (rapid growth, maturation) or pace-of-life syndrome (high metabolic rates, active behavior). Variability across four study species suggests that a combination of different aging and life-history traits conformed with or contradicted the predictions for each species. These findings demonstrate that variation in life span and functional decline among natural populations are linked, genetically underpinned, and can evolve relatively rapidly.
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http://dx.doi.org/10.1111/evo.13127DOI Listing
February 2017

Markers of lipid oxidative damage in the exhaled breath condensate of nano TiO production workers.

Nanotoxicology 2017 02 9;11(1):52-63. Epub 2016 Dec 9.

h UMass Lowell, Department of Public Health , College of Health Sciences , Lowell, MA , USA.

Nanoscale titanium dioxide (nanoTiO) is a commercially important nanomaterial. Animal studies have documented lung injury and inflammation, oxidative stress, cytotoxicity and genotoxicity. Yet, human health data are scarce and quantitative risk assessments and biomonitoring of exposure are lacking. NanoTiO is classified by IARC as a group 2B, possible human carcinogen. In our earlier studies we documented an increase in markers of inflammation, as well as DNA and protein oxidative damage, in exhaled breath condensate (EBC) of workers exposed nanoTiO. This study focuses on biomarkers of lipid oxidation. Several established lipid oxidative markers (malondialdehyde, 4-hydroxy-trans-hexenal, 4-hydroxy-trans-nonenal, 8-isoProstaglandin F2α and aldehydes C-C) were studied in EBC and urine of 34 workers and 45 comparable controls. The median particle number concentration in the production line ranged from 1.98 × 10 to 2.32 × 10 particles/cm with ∼80% of the particles <100 nm in diameter. Mass concentration varied between 0.40 and 0.65 mg/m. All 11 markers of lipid oxidation were elevated in production workers relative to the controls (p < 0.001). A significant dose-dependent association was found between exposure to TiO and markers of lipid oxidation in the EBC. These markers were not elevated in the urine samples. Lipid oxidation in the EBC of workers exposed to (nano)TiO complements our earlier findings on DNA and protein damage. These results are consistent with the oxidative stress hypothesis and suggest lung injury at the molecular level. Further studies should focus on clinical markers of potential disease progression. EBC has reemerged as a sensitive technique for noninvasive monitoring of workers exposed to engineered nanoparticles.
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http://dx.doi.org/10.1080/17435390.2016.1262921DOI Listing
February 2017

Leukotrienes in exhaled breath condensate and fractional exhaled nitric oxide in workers exposed to TiO2 nanoparticles.

J Breath Res 2016 06 30;10(3):036004. Epub 2016 Jun 30.

Department of Occupational Medicine, First Faculty of Medicine, Charles University in Prague and General University Hospital in Prague, Na Bojišti 1, 128 00 Prague 2, Czech Republic.

Human health data regarding exposure to nanoparticles are extremely scarce and biomonitoring of exposure is lacking in spite of rodent pathological experimental data. Potential markers of the health-effects of engineered nanoparticles were examined in 30 workers exposed to TiO2 aerosol, 22 office employees of the same plant, and 45 unexposed controls. Leukotrienes (LT) B4, C4, E4, and D4 were analysed in the exhaled breath condensate (EBC) and urine via liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Fractional exhaled nitric oxide (FeNO) and spirometry was also measured. The median particle number concentration of the aerosol in the production ranged from 1.98  ×  10(4) to 2.32  ×  10(4) particles cm(-3); about 80% of the particles were  <100 nm in diameter. Median total mass concentration varied between 0.4 and 0.65 mg m(-3). All LT levels in workers' EBC were elevated relative to the controls (p  <  0.01). LTs in the EBC sample were correlated with titanium levels. Urinary LTs were not elevated in the workers and office employees. Office workers had higher LTB4 in EBC (p  <  0.05), and higher levels of FeNO (p  <  0.01). FeNO was higher in office employees with allergic diseases and was negatively correlated with smoking (p  <  0.01). In spirometry significant impairment in the workers was seen only for %VCIN and %PEF (both p  <  0.01). Multiple regression analysis confirmed a significant association between production of TiO2 and all cysteinyl LTs in EBC (p  <  0.01) and impaired %VCIN and %PEF (both p  <  0.01). LTB4 was also associated with smoking (p  <  0.01). LT levels complemented our earlier findings of DNA, protein, and lipid damage in the EBC of workers with nanoTiO2 exposures. Cysteinyl LTs in EBC analysis suggest inflammation and potential fibrotic changes in the lungs; they may be helpful for monitoring the biological effect of (nano)TiO2 on workers. Spirometry was not sensitive enough.
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http://dx.doi.org/10.1088/1752-7155/10/3/036004DOI Listing
June 2016

Opposing effects of oxidative challenge and carotenoids on antioxidant status and condition-dependent sexual signalling.

Sci Rep 2016 Mar 22;6:23546. Epub 2016 Mar 22.

Institute of Vertebrate Biology, v.v.i., The Czech Academy of Sciences, Květná 8, Brno 603 65, Czech Republic.

Several recent hypotheses consider oxidative stress to be a primary constraint ensuring honesty of condition-dependent carotenoid-based signalling. The key testable difference between these hypotheses is the assumed importance of carotenoids for redox homeostasis, with carotenoids being either antioxidant, pro-oxidant or unimportant. We tested the role of carotenoids in redox balance and sexual signalling by exposing adult male zebra finches (Taeniopygia guttata) to oxidative challenge (diquat dibromide) and manipulating carotenoid intake. As the current controversy over the importance of carotenoids as antioxidants could stem from the hydrophilic basis of commonly-used antioxidant assays, we used the novel measure of in vivo lipophilic antioxidant capacity. Oxidative challenge reduced beak pigmentation but elicited an increase in antioxidant capacity suggesting resource reallocation from signalling to redox homeostasis. Carotenoids counteracted the effect of oxidative challenge on lipophilic (but not hydrophilic) antioxidant capacity, thereby supporting carotenoid antioxidant function in vivo. This is inconsistent with hypotheses proposing that signalling honesty is maintained through either ROS-induced carotenoid degradation or the pro-oxidant effect of high levels of carotenoid-cleavage products acting as a physiological handicap. Our data further suggest that assessment of lipophilic antioxidant capacity is necessary to fully understand the role of redox processes in ecology and evolution.
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http://dx.doi.org/10.1038/srep23546DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4802301PMC
March 2016

Oxidative stress markers are elevated in exhaled breath condensate of workers exposed to nanoparticles during iron oxide pigment production.

J Breath Res 2016 Feb 1;10(1):016004. Epub 2016 Feb 1.

Charles University in Prague and General University Hospital in Prague, First Faculty of Medicine, Department of Occupational Medicine, Na Bojišti 1, 128 00 Prague 2, Czech Republic.

Markers of oxidative stress and inflammation were analysed in the exhaled breath condensate (EBC) and urine samples of 14 workers (mean age 43  ±  7 years) exposed to iron oxide aerosol for an average of 10  ±  4 years and 14 controls (mean age 39  ±  4 years) by liquid chromatography-electrospray ionization-mass spectrometry/mass spectrometry (LC-ESI-MS/MS) after solid-phase extraction. Aerosol exposure in the workplace was measured by particle size spectrometers, a scanning mobility particle sizer (SMPS) and an aerodynamic particle sizer (APS), and by aerosol concentration monitors, P-TRAK and DustTRAK DRX. Total aerosol concentrations in workplace locations varied greatly in both time and space. The median mass concentration was 0.083 mg m(-3) (IQR 0.063-0.133 mg m(-3)) and the median particle concentration was 66 800 particles cm(-3) (IQR 16,900-86,900 particles cm(-3)). In addition, more than 80% of particles were smaller than 100 nm in diameter. Markers of oxidative stress, malondialdehyde (MDA), 4-hydroxy-trans-hexenale (HHE), 4-hydroxy-trans-nonenale (HNE), 8-isoProstaglandin F2α (8-isoprostane) and aldehydes C6-C12, in addition to markers of nucleic acid oxidation, including 8-hydroxy-2-deoxyguanosine (8-OHdG), 8-hydroxyguanosine (8-OHG), 5-hydroxymethyl uracil (5-OHMeU), and of proteins, such as o-tyrosine (o-Tyr), 3-chlorotyrosine (3-ClTyr), and 3-nitrotyrosine (3-NOTyr) were analysed in EBC and urine by LC-ESI-MS/MS. Almost all markers of lipid, nucleic acid and protein oxidation were elevated in the EBC of workers comparing with control subjects. Elevated markers were MDA, HNE, HHE, C6-C10, 8-isoprostane, 8-OHdG, 8-OHG, 5-OHMeU, 3-ClTyr, 3-NOTyr, o-Tyr (all p  <  0.001), and C11 (p  <  0.05). Only aldehyde C12 and the pH of samples did not differ between groups. Markers in urine were not elevated. These findings suggest the adverse effects of nano iron oxide aerosol exposure and support the utility of oxidative stress biomarkers in EBC. The analysis of urine oxidative stress biomarkers does not support the presence of systemic oxidative stress in iron oxide pigment production workers.
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http://dx.doi.org/10.1088/1752-7155/10/1/016004DOI Listing
February 2016

Ghrelin and endocannabinoids participation in morphine-induced effects in the rat nucleus accumbens.

Psychopharmacology (Berl) 2016 Feb 28;233(3):469-84. Epub 2015 Oct 28.

Department of Organic Technology ICT, Laboratory of Medicinal Diagnostics, Technicka 5, Prague 6, 166 28, Czech Republic.

Rationale And Objectives: In addition to dopamine, endocannabinoids are thought to participate in neural reward mechanisms of opioids. Number of recent studies suggests crucial involvement of ghrelin in some addictive drugs effects. Our previous results showed that ghrelin participates in morphine-induced changes in the mesolimbic dopaminergic system associated with reward processing. The goal of the present study was to test whether the growth hormone secretagogue receptor (GHS-R1A) antagonist JMV2959 was able to influence morphine-induced effects on anandamide (N-arachidonoylethanolamine, AEA) and 2-arachidonoylglycerol (2-AG) in the nucleus accumbens shell (NACSh).

Methods: We used in vivo microdialysis to determine changes in levels of AEA and 2-AG in the NACSh in rats following (i) an acute morphine dose (5, 10 mg/kg s.c.) with and without JMV2959 pretreatment (3, 6 mg/kg i.p.) or (ii) a morphine challenge dose (5 mg/kg s.c.) with and without JMV2959 (3, 6 mg/kg i.p.) pretreatment, administered during abstinence following repeated doses of morphine (5 days, 10-40 mg/kg). Co-administration of ghrelin (40 ug/kg i.p.) was used to verify the ghrelin mechanisms involvement.

Results: Pretreatment with JMV2959 significantly and dose-dependently reversed morphine-induced anandamide increases in the NACSh in both the acute and longer-term models, resulting in a significant AEA decrease. JMV2959 significantly intensified acute morphine-induced decreases in accumbens 2-AG levels and attenuated morphine challenge-induced 2-AG decreases. JMV2959 pretreatment significantly reduced concurrent morphine challenge-induced behavioral sensitization. JMV2959 pretreatment effects were abolished by co-administration of ghrelin.

Conclusions: Our results indicate significant involvement of ghrelin signaling in morphine-induced endocannabinoid changes in the NACSh.
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http://dx.doi.org/10.1007/s00213-015-4119-3DOI Listing
February 2016

Reward related neurotransmitter changes in a model of depression: An in vivo microdialysis study.

World J Biol Psychiatry 2015 Oct 7;16(7):521-35. Epub 2015 Oct 7.

c Department of Pharmacology , Third Faculty of Medicine, Charles University , Prague , Czech Republic , and.

Objectives: The self-medication hypothesis assumes that symptoms related to potential monoaminergic deficits in depression may be relieved by drug abuse. The aim of this study was to elucidate the neurotransmitter changes in a rat model of depression by measuring their levels in the nucleus accumbens shell, which is typically involved in the drug of abuse acquisition mechanism.

Methods: Depression was modelled by the olfactory bulbectomy (OBX) in Wistar male rats. In vivo microdialysis was performed, starting from the baseline and following after a single methamphetamine injection and behaviour was monitored. The determination of neurotransmitters and their metabolites was performed by high-performance liquid chromatography combined with mass spectrometry.

Results: OBX animals had lower basal levels of dopamine and serotonin and their metabolites. However, γ-aminobutyric acid (GABA) and glutamate levels were increased. The methamphetamine injection induced stronger dopamine and serotonin release in the OBX rats and lower release of glutamate in comparison with sham-operated rats; GABA levels did not differ significantly.

Conclusions: This study provides an evidence of mesolimbic neurotransmitter changes in the rat model of depression which may elucidate mechanisms underlying intravenous self-administration studies in which OBX rats were demonstrated to have higher drug intake in comparison to intact controls.
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http://dx.doi.org/10.3109/15622975.2015.1077991DOI Listing
October 2015

Biochemical, histopathological and morphological profiling of a rat model of early immune stimulation: relation to psychopathology.

PLoS One 2015 20;10(1):e0115439. Epub 2015 Jan 20.

Prague Psychiatric Center, Prague, Czech Republic; National Institute of Mental Health, Klecany, Czech Republic; Third Faculty of Medicine, Charles University, Prague, Czech Republic.

Perinatal immune challenge leads to neurodevelopmental dysfunction, permanent immune dysregulation and abnormal behaviour, which have been shown to have translational validity to findings in human neuropsychiatric disorders (e.g. schizophrenia, mood and anxiety disorders, autism, Parkinson's disease and Alzheimer's disease). The aim of this animal study was to elucidate the influence of early immune stimulation triggered by systemic postnatal lipopolysaccharide administration on biochemical, histopathological and morphological measures, which may be relevant to the neurobiology of human psychopathology. In the present study of adult male Wistar rats we examined the brain and plasma levels of monoamines (dopamine, serotonin), their metabolites, the levels of the main excitatory and inhibitory neurotransmitters glutamate and γ-aminobutyric acid and the levels of tryptophan and its metabolites from the kynurenine catabolic pathway. Further, we focused on histopathological and morphological markers related to pathogenesis of brain diseases--glial cell activation, neurodegeneration, hippocampal volume reduction and dopaminergic synthesis in the substantia nigra. Our results show that early immune stimulation in adult animals alters the levels of neurotransmitters and their metabolites, activates the kynurenine pathway of tryptophan metabolism and leads to astrogliosis, hippocampal volume reduction and a decrease of tyrosine hydroxylase immunoreactivity in the substantia nigra. These findings support the crucial pathophysiological role of early immune stimulation in the above mentioned neuropsychiatric disorders.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0115439PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4300081PMC
September 2015

Multimarker screening of oxidative stress in aging.

Oxid Med Cell Longev 2014 16;2014:562860. Epub 2014 Jul 16.

Institute of Chemical Technology, Technicka 5, 166 28 Prague 6, Czech Republic.

Aging is a complex process of organism decline in physiological functions. There is no clear theory explaining this phenomenon, but the most accepted one is the oxidative stress theory of aging. Biomarkers of oxidative stress, substances, which are formed during oxidative damage of phospholipids, proteins, and nucleic acids, are present in body fluids of diseased people as well as the healthy ones (in a physiological concentration). 8-iso prostaglandin F2α is the most prominent biomarker of phospholipid oxidative damage, o-tyrosine, 3-chlorotyrosine, and 3-nitrotyrosine are biomarkers of protein oxidative damage, and 8-hydroxy-2(')-deoxyguanosine and 8-hydroxyguanosine are biomarkers of oxidative damage of nucleic acids. It is thought that the concentration of biomarkers increases as the age of people increases. However, the concentration of biomarkers in body fluids is very low and, therefore, it is necessary to use a sensitive analytical method. A combination of HPLC and MS was chosen to determine biomarker concentration in three groups of healthy people of a different age (twenty, forty, and sixty years) in order to find a difference among the groups.
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http://dx.doi.org/10.1155/2014/562860DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4124763PMC
March 2015

Sex differences in methamphetamine pharmacokinetics in adult rats and its transfer to pups through the placental membrane and breast milk.

Drug Alcohol Depend 2014 Jun 29;139:138-44. Epub 2014 Mar 29.

Charles University in Prague, Third Faculty of Medicine, Department of Normal, Pathological and Clinical Physiology, Ke Karlovu 4, 120 00 Prague, Czech Republic.

Background: Methamphetamine (METH) abuse is a growing health problem worldwide, and METH use during pregnancy not only endangers the mother's health but also the developing fetus. To provide better insight into these risks, we performed the following experiments.

Method: First, we investigated how sex influences the pharmacokinetics of METH and amphetamine (AMP) in male and female rats. Subsequently, we simulated chronic exposure of prenatal infants to METH abuse by investigating brain and plasma levels of METH and AMP in dams and pups. Finally, we modeled chronic exposure of infants to METH via breast milk and investigated sex differences in pups with regard to drug levels and possible sensitization effect of chronic prenatal METH co-treatment.

Results: We observed significantly higher levels of METH and AMP in the plasma and brain of female rats compared to males. Additionally, brain concentrations of METH and AMP in pups exposed to METH prenatally were equivalent to 62.13% and 37.78% relative to dam, respectively. Plasma concentrations of AMP where equivalent to 100% of the concentration in dams, while METH was equivalent to only 36.98%. Finally, we did not observe a significant effect relative to sex with regard to METH/AMP levels or sensitization effects linked to prenatal METH exposure.

Conclusion: We demonstrated that female rats display higher levels of METH and AMP, thus indicating a greater risk of addiction and toxicity. Furthermore, our data show that pups are exposed to both METH and AMP following dam exposure.
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http://dx.doi.org/10.1016/j.drugalcdep.2014.03.023DOI Listing
June 2014

Occupational asthma follow-up--which markers are elevated in exhaled breath condensate and plasma?

Int J Occup Med Environ Health 2014 Apr 19;27(2):206-15. Epub 2014 Mar 19.

Department of Occupational Medicine of the First Faculty of Medicine, Charles University in Prague and General University Hospital, Prague, Czech Republic,

Objectives: To search for optimal markers in the exhaled breath condensate (EBC), plasma and urine that would reflect the activity/severity of occupational asthma (OA) after the withdrawal from the exposure to the allergen.

Material And Methods: Markers of oxidative stress: 8-iso-prostaglandin F2α (8-isoprostane, 8-ISO), malondialdehyde (MDA), 4-hydroxy-trans-2-nonenale (HNE), cysteinyl leukotrienes (LT) and LTB4 were determined using liquid chromatography and mass spectrometry in 43 subjects with immunological OA (49.3 ± 11.8 years), removed from the exposure to the sensitizing agent 10.5 ± 6.5 years ago; and in 20 healthy subjects (49.0 ± 14.9 years). EBC was harvested both before and after the methacholine challenge test. In parallel, identical markers were collected in plasma and urine. The results were analyzed together with forced expiratory volume in one second (FEV1), blood eosinophils, immunoglobulin E (IgE) and eosinophilic cationic protein (ECP) and statistically evaluated (Spearman rank correlation rS, two- or one-sample t tests and alternatively Kruskal Wallis or pair Wilcoxon tests).

Results: Several parameters of lung functions were lower in the patients (FEV1% predicted, MEF25% and MEF50%, Rtot%, p < 0.001). Shorter time interval since the removal from the allergen exposure correlated with higher ECP (rS = 0.375) and lower FEV1%, MEF25% and MEF50% after methacholine challenge (rS = -0.404, -0.425 and -0.532, respectively). In the patients, IgE (p < 0.001) and ECP (p = 0.009) was increased compared to controls. In EBC, 8-ISO and cysteinyl LTs were elevated in the asthmatics initially and after the challenge. Initial 8-ISO in plasma correlated negatively with FEV1 (rS = -0.409) and with methacholine PD20 (rS = -0.474). 8-ISO in plasma after the challenge correlated with IgE (rS = 0.396).

Conclusions: The improvement in OA is very slow and objective impairments persist years after removal from the exposure. Cysteinyl LTs and 8-ISO in EBC and 8-ISO in plasma might enrich the spectrum of useful objective tests for the follow-up of OA.
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http://dx.doi.org/10.2478/s13382-014-0243-2DOI Listing
April 2014

Immunomagnetic molecular probe with UHPLC-MS/MS: a promising way for reliable bronchial asthma diagnostics based on quantification of cysteinyl leukotrienes.

J Pharm Biomed Anal 2013 Jul-Aug;81-82:108-17. Epub 2013 Apr 8.

Institute of Chemical Technology, Technicka 5, 166 28 Prague 6, Czech Republic.

A sensitive and precise method for simultaneous quantification of cysteinyl leukotrienes (=cys LTs) - leukotriene C4 (=LTC4), leukotriene D4 (=LTD4) and leukotriene E4 (=LTE4) - essential biomarkers of bronchial asthma present in exhaled breath condensate (=EBC) was developed. An immunomagnetic molecular probe was prepared by anchoring cysteinyl leukotrienes antibody on the surface of functionalized monodispersed magnetic particles and used to selectively isolate cys LTs from biological matrices - EBC, plasma and urine. Immobilization and the immunoaffinity capture procedures were optimized to maximize the amount of separated cys LTs, which were detected "off-beads" after acidic elution by UHPLC-ESI-MS/MS operated in a multiple reaction monitoring mode. The developed method was characterized with high precision ≤13.6% (intra-day precision determined as RSD) and ≤14.5% (inter-day precision determined as RSD), acceptable accuracy ≤18.5% (determined as RE), and high recovery of immunoseparation (≥93.1%) in aforementioned biological matrices. The applicability of the method was demonstrated on EBC, plasma and urine clinical samples of patients with various subtypes of bronchial asthma (occupational, steroid-resistant, moderate with and without corticosteroids therapy) and healthy subjects where reasonable differences in cys LTs concentration levels were found. Combining extremely selective immunomagnetic separation with highly sensitive and precise detection step, the developed method was used to aid diagnosis, predict the most effective therapy, and monitor the response to treatment. The detection of elevated inflammatory mediators (cys LTs) in EBC of subjects with relatively asymptomatic asthma and normal pulmonary function tests could offer a novel way for monitoring the lung inflammation and perhaps initiating treatment in an earlier stage.
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http://dx.doi.org/10.1016/j.jpba.2013.03.026DOI Listing
December 2013

Behavioral, neurochemical and pharmaco-EEG profiles of the psychedelic drug 4-bromo-2,5-dimethoxyphenethylamine (2C-B) in rats.

Psychopharmacology (Berl) 2013 Jan 29;225(1):75-93. Epub 2012 Jul 29.

Prague Psychiatric Center, Ústavní 91, 181 03 Bohnice, Prague 8, Czech Republic.

Rationale And Objectives: Behavioral, neurochemical and pharmaco-EEG profiles of a new synthetic drug 4-bromo-2,5-dimethoxyphenethylamine (2C-B) in rats were examined.

Materials And Methods: Locomotor effects, prepulse inhibition (PPI) of acoustic startle reaction (ASR), dopamine and its metabolite levels in nucleus accumbens (NAc), EEG power spectra and coherence in freely moving rats were analysed. Amphetamine was used as a reference compound.

Results: 2C-B had a biphasic effect on locomotion with initial inhibitory followed by excitatory effect; amphetamine induced only hyperlocomotion. Both drugs induced deficits in the PPI; however they had opposite effects on ASR. 2C-B increased dopamine but decreased 3,4-dihydroxyphenylacetic acid (DOPAC) in the NAc. Low doses of 2C-B induced a decrease in EEG power spectra and coherence. On the contrary, high dose of 2C-B 50 mg/kg had a temporally biphasic effect with an initial decrease followed by an increase in EEG power; decrease as well as increase in EEG coherence was observed. Amphetamine mainly induced an increase in EEG power and coherence in theta and alpha bands. Increases in the theta and alpha power and coherence in 2C-B and amphetamine were temporally linked to an increase in locomotor activity and DA levels in NAc.

Conclusions: 2C-B is a centrally active compound similar to other hallucinogens, entactogens and stimulants. Increased dopamine and decreased DOPAC in the NAc may reflect its psychotomimetic and addictive potential and monoaminoxidase inhibition. Alterations in brain functional connectivity reflected the behavioral and neurochemical changes produced by the drug; a correlation between EEG changes and locomotor behavior was observed.
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http://dx.doi.org/10.1007/s00213-012-2797-7DOI Listing
January 2013

Leukotrienes B4, C4, D4 and E4 in the exhaled breath condensate (EBC), blood and urine in patients with pneumoconiosis.

Ind Health 2012 13;50(4):299-306. Epub 2012 Jun 13.

Department of Occupational Medicine of the 1st Faculty of Medicine, Charles University in Prague, Czech Republic.

Leukotrienes (LTs) are involved in the pathogenesis of lung fibrosis and were increased in exhaled breath condensate (EBC) of the patients with pneumoconiosis. However the possible influence of extra-pulmonary disorders on the EBC markers is not known. Therefore in parallel with EBC, LTs' levels in the plasma and urine were measured in patients with pneumoconiosis (45 × asbestos exposure, 37 × silica exposure) and in 27 controls. Individual LTs B4, C4, D4 and E4 were measured by liquid chromatography - electrospray ionization - tandem mass spectrometry (LC-ESI-MS/MS). In EBC, LT D4 and LT E4 were increased in both groups of patients (p<0.001 and p<0.05), comparing with the controls. Both LT B4 and cysteinyl LTs were elevated in asbestos-exposed subjects (p<0.05). Asbestosis with more severe radiological signs (s1/s2-t3/u2) and lung functions impairment has shown higher cysteinyl LTs and LT C4 in the EBC (p<0.05) than mild asbestosis (s1/s0-s1/s1). In addition, in the subjects with asbestosis, cysteinyl LTs in EBC correlated with TLC (-0.313, p<0.05) and TLCO/Hb (-0.307, p<0.05), and LT C4 with TLC (-0.358, p<0.05). In pneumoconioses, EBC appears the most useful from the 3 fluids studied.
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http://dx.doi.org/10.2486/indhealth.ms1274DOI Listing
February 2013

Increased oxidative/nitrosative stress markers measured non- invasively in patients with high 2,3,7,8-tetrachloro-dibenzo-p-dioxin plasma level.

Neuro Endocrinol Lett 2011 ;32 Suppl 1:71-6

Department of Occupational Medicine of the First Faculty of Medicine and General University Hospital, Charles University Prague, Czech Republic.

Objectives: 2,3,7,8-tetrachloro-dibenzo-p-dioxin (TCDD) is a highly toxic persistent environmental contaminant, classified as a human carcinogen affecting any target organ. The mechanism of carcinogenesis by TCDD is unclear as TCDD shows a lack of direct genotoxicity. Experimental studies also support the role of oxidative stress in TCDD neurotoxicity and vascular dysfunction. The aim was to investigate markers of oxidative/nitrosative stress and inflammation using non-invasive methods in subjects who got ill due to severe occupational exposure to TCDD in the years 1965-1968.

Methods: In 11 TCDD-exposed patients, and 16 controls, the analysis of following oxidative products of lipids, proteins and nucleic acids in plasma, urine and exhaled breath condensate (EBC) was performed: 8-iso-prostaglandin F2α (8-isoprostane), 4-hydroxy-trans-2-nonenale (HNE), malondialdehyde (MDA), o-tyrosine (o-Tyr), 8-hydroxyguanosine (8-OHG), 8-hydroxy-2´-deoxy-guanosine (8-OHdG), 5-hydroxymethyluracil (5-OHMeU). In addition, nitric-oxide-tyrosine (NO-Tyr) and leukotriene (LT) B4, C4, D4, and E4 were detected by liquid chromatography-mass spectrometry/mass spectrometry (LC-ESI-MS/MS). TCDD was measured by HRGC/HRMS, body lipid content by densitometry. Single-photon emission spectrometry (SPECT) of the brain was performed and compared with the findings of the patients in 2008.

Results: Mean TCDD plasma level in 2010 was 175 ± 162 pg/g lipids (population level about 2 pg/g), total TCDD content in the body 5.16 ± 4.62 mg. Reduction of cerebral blood flow in SPECT progressed in 8 patients, finding was stable in 2 subjects, and improvement occurred in 1 patient. In the EBC, 10 from 12 markers (all except LT D4 and LT E4), were significantly increased in the patients (p<0.05). In the urine, 7 markers were significantly higher than in the controls (p<0.05): 8-isoprostane, MDA, HNE, LT C4, LT E4, o-Tyr and NO-Tyr. In plasma, only NO-Tyr and 8-OHG were elevated (p<0.05).

Conclusion: NO-Tyr was increased in all matrices in dioxin-exposed patients. EBC is not limited to lung disorders as the markers of oxidative stress and inflammation were elevated in EBC of patients with normal lung functions. TCDD-induced oxidative stress and inflammation markers can be detected non-invasively in the EBC and urine in the follow-up of the highly-exposed patients. Their prognostic value, however, needs to be elucidated.
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April 2012

Oxidative stress markers in exhaled breath condensate in lung fibroses are not significantly affected by systemic diseases.

Ind Health 2011 20;49(6):746-54. Epub 2011 Oct 20.

Charles University in Prague, 1st Faculty of Medicine, Department of Occupational Medicine of the 1st Faculty of Medicine, Prague, Czech Republic.

Exhaled breath condensate (EBC) is assumed to reflect processes in the lungs, yet it is unknown whether oxidative stress markers in EBC are affected by systemic disorders (atherosclerosis, hypertension, diabetes) or whether lung diseases increase markers in plasma and urine. 8-isoprostane, 4-hydroxy-trans-2-nonenale (HNE) and malondialdehyde (MDA) were measured using liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS/MS) in EBC, plasma and urine in 82 patients (45 with asbestosis and hyalinosis, and 37 with silicosis) and in 29 control subjects. 8-isoprostane and HNE in EBC, and HNE in urine were higher in both groups of patients. In addition, 8-isoprostane in plasma and urine, and MDA in urine were higher in asbestos-exposed patients and MDA in plasma in silicotics, with this marker in plasma correlated with the grade of silicosis. In all subjects, 8-isoprostane in EBC correlated with urine (r=0.38, p<0.001) and plasma levels (r=0.28, p=0.003), and HNE and MDA with urine levels (r=0.31, p<0.001; r=0.23, p=0.016, respectively). Most markers positively correlated with lung function impairment, EBC markers negatively with vitamin E supplementation. To conclude: The influence of satisfactorily controlled systemic disorders on markers in EBC in patients with pneumoconioses is not significant. In addition to oxidative stress markers in EBC, lung fibroses may increase oxidative stress markers in plasma and urine.
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http://dx.doi.org/10.2486/indhealth.ms1237DOI Listing
April 2012

Determination of cysteinyl leukotrienes in exhaled breath condensate: method combining immunoseparation with LC-ESI-MS/MS.

J Chromatogr B Analyt Technol Biomed Life Sci 2011 Aug 13;879(23):2220-8. Epub 2011 Jun 13.

Institute of Chemical Technology, Technická 5, 166 28 Prague 6, Czech Republic.

A rapid and precise method for the identification and quantification of cysteinyl leukotrienes (leukotriene C(4), leukotriene D(4) and leukotriene E(4)), essential markers of bronchial asthma, in exhaled breath condensate was developed. The protocol consists of immunoaffinity separation and a detection step, liquid chromatography combined with electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). In particular, the selected reaction monitoring mode was used for its extremely high degree of selectivity and the stable-isotope-dilution assay for its high precision of quantification. The developed method was characterized with a high precision (≤ 7.7%, determined as RSD), an acceptable accuracy (90.4-93.7%, determined as recovery), a low limit of detection (≤ 2 pg/ml EBC) and a low limit of quantification (≤ 10 pg/ml EBC). It was compared to other simple, clinically appropriate combinations of pre-treatment methods (solid phase extraction and lyophilization) with LC/MS. Finally, the method (a combination of immunoaffinity separation with LC-MS) was successfully tested in a clinical study where a significant difference was found in the concentration levels of cysteinyl leukotrienes between patients with occupational bronchial asthma and healthy subjects.
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http://dx.doi.org/10.1016/j.jchromb.2011.06.004DOI Listing
August 2011

Monitoring of dopamine and its metabolites in brain microdialysates: method combining freeze-drying with liquid chromatography-tandem mass spectrometry.

J Chromatogr A 2011 May 26;1218(21):3382-91. Epub 2011 Feb 26.

Institute of Chemical Technology, Technicka 5, Prague 6, Czech Republic.

A sensitive assay method was developed for a parallel, rapid and precise determination of dopamine and its metabolites, homovanillic acid, 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid, from brain microdialysates. The method consisted of a pre-treatment step, freeze-drying (lyophilization), to concentrate dopamine and its metabolites from the microdialysates, and a detection step using liquid chromatography combined with electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). In particular, the reaction monitoring mode was selected for its extremely high degree of selectivity and the stable-isotope-dilution assay for its high precision of quantification. The developed method was characterized by the following parameters: the precision of the developed method was determined as ≥88.6% for dopamine, ≥89.9% for homovanillic acid, ≥86.1% for 3-methoxytyramine and ≥88.1% for 3,4-dihydroxyphenylacetic acid; the mean accuracy was determined as ≥88.2% for dopamine, ≥88.3% for homovanillic acid, ≥85.9% for 3-methoxytyramine and ≥88.6% for 3,4-dihydroxyphenylacetic acid. The developed method was compared to (1) other combinations of pre-treatment methods (solid phase extraction and nitrogen stripping) with LC-MS and (2) another detection method, liquid chromatography, with electrochemical detection. The novel developed method using combination of lyophilization with LC-ESI-MS/MS was tested on real samples obtained from the nucleus accumbens of rat pups after an acute methamphetamine administration. It was proven that the developed assay could be applied to both a simultaneous analysis of all four substrates (dopamine, homovanillic acid, 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid) in microdialysis samples acquired from the rat brain and the monitoring of their slight concentration changes on a picogram level over time following methamphetamine stimulus.
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http://dx.doi.org/10.1016/j.chroma.2011.02.006DOI Listing
May 2011

Cellular and behavioural effects of a new steroidal inhibitor of the N-methyl-d-aspartate receptor 3α5β-pregnanolone glutamate.

Neuropharmacology 2011 Jul-Aug;61(1-2):61-8. Epub 2011 Feb 24.

Institute of Physiology, Academy of Sciences of the Czech Republic, Videnska 1083, Prague 4, Czech Republic.

Preclinical studies have demonstrated a considerable role for N-methyl-d-aspartate (NMDA) receptors in excitotoxicity and the concurrent neuroprotective effect of NMDA receptor antagonists. Because NMDA receptors are one of the most widespread receptors in the central nervous system, application of their antagonist often leads to serious side effects ranging from motor impairment to induction of schizophrenic-like psychosis. Therefore, we have initiated development and testing of a novel synthetic NMDA receptor antagonist derived from naturally occurring neurosteroids. 20-oxo-5β-pregnan-3α-yl-l-glutamyl-1-ester (3α5βP-Glu) is a novel synthetic steroidal inhibitor of the NMDA receptor. Our results show that 3α5βP-Glu preferentially inhibits tonically activated NMDA receptors, is able to cross the blood brain barrier, does not induce psychotomimetic symptoms (such as hyperlocomotion and sensorimotor gating deficit) and reduced an excitotoxic damage of brain tissue and subsequent behavioural impairment in rats. In particular, 3α5βP-Glu significantly ameliorated neuronal damage in the dentate gyrus and subiculum, and improved behavioural performance in active allothetic place avoidance tasks (AAPA, also known as the carousel maze) after bilateral NMDA-induced lesions to the hippocampi. These findings provide a possible new therapeutic approach for the treatment of diseases induced by NMDA receptor overactivation.
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http://dx.doi.org/10.1016/j.neuropharm.2011.02.018DOI Listing
April 2012

Prenatal methamphetamine exposure affects the mesolimbic dopaminergic system and behavior in adult offspring.

Int J Dev Neurosci 2009 Oct 8;27(6):525-30. Epub 2009 Jul 8.

Prague Psychiatric Center, Department of Biochemistry and Brain Pathophysiology, Ustavní 91, 181 03 Prague 8, Bohnice, Czech Republic.

Methamphetamine is a commonly abused psychostimulant that causes addiction and is often abused by pregnant women. Acute or chronic administration of methamphetamine elevates the levels of the extracellular monoamine neurotransmitters, such as dopamine. The aim of the present study was to show whether prenatal exposure to methamphetamine (5mg/kg, entire gestation) or saline in Wistar rats induces changes in dopamine levels and its metabolites in the nucleus accumbens, and in behavior (locomotor activity, rearing, and immobility) after the administration of a challenge dose of methamphetamine (1mg/kg) or saline in male offspring. We found that adult offspring prenatally exposed to methamphetamine had higher basal levels of dopamine (about 288%), dihydroxyphenylacetic acid (about 67%) and homovanillic acid (about 74%) in nucleus accumbens. An increased basal level of dopamine corresponds to lower basal immobility in offspring prenatally exposed to methamphetamine. The acute injection of methamphetamine in adulthood increased the level of dopamine in the nucleus accumbens, which is related to an increase of locomotion and rearing (exploration). In addition, prenatally methamphetamine-exposed rats showed higher response to the challenge dose of methamphetamine, when compared to prenatally saline-exposed rats. In conclusion, rats exposed to methamphetamine in utero have shown changes in the mesolimbic dopaminergic system and were more sensitive to the administration of the acute dose of methamphetamine in adulthood.
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http://dx.doi.org/10.1016/j.ijdevneu.2009.06.012DOI Listing
October 2009

Rapid and easy method for monitoring oxidative stress markers in body fluids of patients with asbestos or silica-induced lung diseases.

J Chromatogr B Analyt Technol Biomed Life Sci 2009 Aug 13;877(24):2477-86. Epub 2009 Jun 13.

Institute of Chemical Technology, Technická 5, 166 28 Prague 6, Czech Republic.

Sensitive assay method was developed for a parallel, rapid and precise determination of the most prominent oxidative stress biomarkers: 8-iso-prostaglandin F(2alpha), malondialdehyde and 4-hydroxynonenal. The method consisted of a pre-treatment part a solid-phase extraction, for rapid and effective isolation of biomarkers from body fluids (exhaled breath condensate, plasma and urine) and the detection method LC-ESI-MS/MS, where the selected reaction monitoring mode was used for its extremely high degree of selectivity and the stable-isotope-dilution assay for its high precision of quantification. The developed method was characterized by the following parameters: the imprecision was below 14.3%, the mean inaccuracy was determined to be lower than 13.1%. The method was tested on samples obtained from patients diagnosed with asbestosis, pleural hyalinosis or silicosis, i.e. occupational lung diseases caused by fibrogenic dusts, inducing oxidative stress in the respiratory system, and then compared to samples from healthy subjects. The difference in concentration levels of biomarkers between the two groups was perceptible in all the body fluids (the difference observed in an exhaled breath condensate was statistically most significant).
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http://dx.doi.org/10.1016/j.jchromb.2009.06.008DOI Listing
August 2009

Determination of 8-iso-prostaglandin F(2alpha) in exhaled breath condensate using combination of immunoseparation and LC-ESI-MS/MS.

J Chromatogr B Analyt Technol Biomed Life Sci 2008 May 29;867(1):8-14. Epub 2008 Feb 29.

Institute of Chemical Technology, Technická 5, 166 28 Prague 6, Czech Republic.

Rapid and precise method for the determination of 8-iso-prostaglandin F(2alpha), an essential marker of the oxidative stress, in exhaled breath condensate (EBC) was developed. The protocol consisted of stable isotope dilution, immunoseparation combined with selective and sensitive LC-ESI-MS/MS operated in multiple reaction monitoring (MRM) mode. The imprecision of the developed method was below 8.8%, the parameter of mean inaccuracy was determined as <9.6% (0-250pg of 8-iso-prostaglandin F(2alpha)/ml EBC). The limit of detection (LOD) was 1 pg/ml EBC and limit of quantification (LOQ) 5 pg/ml EBC. A significant difference in 8-iso-prostaglandin F(2alpha) content between the group of asbestosis patients and healthy volunteers was found.
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http://dx.doi.org/10.1016/j.jchromb.2008.02.019DOI Listing
May 2008