Effects of ginkgo biloba extract on cation currents in rat ventricular myocytes

Ginkgo biloba extract (GBE), a valuable natural product for cerebral and cardiovascular diseases, is mainly composed of two classes of constituents: terpene lactones (e.g., ginkgolide A and B, bilobalide) and flavone glycosides (e.g., quercetin and kaempferol). Its electrophysiological action in heart is yet unclear. In the present study, using whole-cell patch clamp technique, we investigated electrophysiological effects of GBE on cation channel currents in ventricular myocytes isolated from rat hearts. We found that GBE 0.01-0.1% inhibited significantly the sodium current (I-Na), L-type calcium current (I-Ca) and transient outward potassium current (IKto) in a concentration-dependent manner. Surprisingly, its main ingredients, ginkgolide A (GB A), ginkgolide B (GB B) and bilobalide (GB BA) at 0.1 mM did not exhibit any significant effect on these cation channel currents. These results suggested that GBE is a potent non-selective cation channel modulator in cardiaomyocytes. Other constituents (rather than GB A, GB B and GB BA) might be responsible for the observed inhibitory effects of GBE on cation channels. (C) 2004 Elsevier Inc. All rights reserved.

The profound effects of microcystin on cardiac antioxidant enzymes, mitochondrial function and cardiac toxicity in rat

Deaths from microcystin toxication have widely been attributed to hypovolemic shock due to hepatic interstitial hemorrhage, while some recent studies suggest that cardiogenic complication is also involved. So far, information on cardiotoxic effects of MC has been rare and the underlying mechanism is still puzzling. The present study examined toxic effects of microcystins on heart muscle of rats intravenously injected with extracted MC at two doses, 0.16LD(50) (14 mu g MC-LReq kg(-1) body weight) and 1LD(50) (87 mu g MC-LReq kg(-1) body weight). In the dead rats, both TTC staining and maximum elevations of troponin I levels confirmed myocardial infarction after MC exposure, besides a serious interstitial hemorrhage in liver. In the 1LD(50) dose group, the coincident falls in heart rate and blood pressure were related to mitochondria dysfunction in heart, while increases in creatine kinase and troponin I levels indicated cardiac cell injury. The corresponding pathological alterations were mainly characterized as loss of adherence between cardiac myocytes and swollen or ruptured mitochondria at the ultrastructural level. MC administration at a dose of 1LD(50) not only enhanced activities and up-regulated mRNA transcription levels of antioxidant enzymes, but also increased GSH content. At both doses, level of lipid peroxides increased obviously, suggesting serious oxidative stress in mitochondria. Simultaneously. complex I and III were significantly inhibited, indicating blocks in electron flow along the mitochondrial respiratory chain in heart. In conclusion, the findings of this study implicate a role for MC-induced cardiotoxicity as a potential factor that should be considered when evaluating the mechanisms of death associated with microcystin intoxication in Brazil. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Distribution of microcystins in various organs (heart, liver, intestine, gonad, brain, kidney and lung) of Wistar rat via intravenous injection

The distribution of microcystins (MCs) in various tissues of Wistar rats was studied under laboratory conditions. Rats were injected intravenously (i.v.) with extracted MCs at a dose of 80 mu g MC-LRequivalent/kg body weight. MCs concentrations in various tissues were detected at 1, 2. 4, 6, 12 and 24 h post-injection using liquid chromatography-mass spectrometry (LC-MS). The highest concentration of MCs was found in kidney (0.034-0.295 mu g/g dry weight), followed by lung (0.007-0.067 mu g/g dry weight), stomach (0.010-0.058 mu g/g dry weight) and liver (0.003-0.052 mu g/g dry weight). The maximum MCs content in the whole body of rat, 2.9% of the injected dose, was observed at 2 h post-injection. MCs concentration was higher in kidney than in liver during the experiment, and two peaks of MCs concentration (at 2 and 24 h, respectively) were observed in kidney, indicating that MCs can be excreted directly via kidney of rat. Though heart, intestine, spleen, brain, gonad and stomach contained less than 0.2% of injected MCs during the whole experiment stage, the presence of MCs in these tissues represents potential damage to them. (c) 2008 Elsevier Ltd. All Fights reserved.

Quantitative Determination of Microcystins in Rat Plasma by LC-ESI Tandem MS

A rapid and sensitive method was developed and validated for the determination of MCYST (microcystin)-RR, -LR, and [Dha(7)] MCYST-LR in rat plasma by liquid chromatography-tandem mass spectrometry. The analytes were extracted from rat plasma by protein precipitation, followed by solid-phase extraction. Liquid chromatography with electrospray ionization mass spectrometry, operating in selected reaction monitoring (SRM) mode, was used to quantify MCYST-RR, -LR, and [Dha(7)] MCYST-LR in rat plasma. The recoveries for each analyte in rat plasma ranged from 70.8 to 88.7%. The calibration curve was linear within the range from 0.005 to 1.25 mu g mL(-1). The limit of detection were 1.4, 1.0, 0.6 ng mL(-1) for MCYST-RR, -LR, and [Dha(7)] MCYST-LR. The overall precision was determined on three different days. The values for within- and between-day precision in rat plasma were within 15%. This method was applied to the identification and quantification of microcystins in rat plasma with acute exposure of microcystins via intravenous injection.

Induction of hepatic cytochrome P4501A1/2B activity and disruption of thyroglobulin synthesis/secretion by mono-ortho polychlorinated biphenyl and its hydroxylated metabolites in rat cell lines

As the active metabolites of polychlorinated biphenyl (PCBs), hydroxylated polychlorinated biphenyls (OH-PCBs) are found in wildlife and human tissues. They have been proposed as main contributors for endocrine disruption of PCBs in living organisms. In this study, mono-ortho PCB 156 and its hydroxylated metabolites 4'-OH-PCB 159, 4'-OH-PCB 121, and 4'-OH-PCB 72 were selected to investigate the toxic effects on rat hepatoma H4IIE cell line and rat thyroid follicle FRTL-5 cell line at concentrations of 1, 10(2), 10(4) nM. 7-Ethoxyresorufin-O-deethylase (EROD) and 7-pentoxyresorufin-O-dealkylase (PROD) activities were determined with micro-EROD/PROD to indicate cytochrome P4501 A1 (CYP1A1) and cytochrome P4502B (CYP2B) induction in the H4IIE cell after exposure for 72 h. To assess thyroid disruption of these compounds, thyroglobulin concentrations also were detected inside FRTL-5 cell with immunocellularchemistry and in its medium with radioimmunoassay after exposure for 24 It. Significant inductions of EROD activity by PCB 156 at 102 and 104 nM (p < 0.05) were observed, but no effects by the three OH-PCBs in H4IIE cell line. 7-Pentoxyresorufin-O-dealkylase activities were induced only by 10(4) nM of PCB156 and the three OH-PCBs (p < 0.05). Meanwhile, significant increases of thyroglobulin concentrations were observed in the medium of FRTL-5 cell exposed to 4'-OH-PCB 121 and 4'-OH-PCB 72 at all of the test concentrations (p < 0.05), but not to the other compounds. The results demonstrated that mono-ortho PCBs mainly could be metabolized to hydroxylated metabolites through CYP1A1 instead of CYP2B. Moreover, after being metabolized, OH-PCBs still sustained the ability to induce PROD activity and did exhibit the disruption on thyroglobulin synthesis/excretion in rat cells.

Evaluation of bioeffects of a long-term administering ChangLe on rat by proton NMR method and biochemical examination

High resolution H-1 nuclear magnetic resonance ( NMR) spectroscopy has been employed to assess long-term toxicological effects of ChangLe (a kind of rare earth complex applied in agriculture). Male Wistar rats were administrated orally with ChangLe at doses of 0, 0.1, 0.2, 2.0, 10 and 20 mg/kg body weight daily, respectively, for 6 months. Urine was collected at-day 30, 60, go and serum samples were taken after 6 months. Many low-molecular weight metabolites were identified by H-1 NMR spectra of rat urine. A decrease in citrate and an increase in ketone bodies, creatinine, DMA, DMG, TMAO, and taurine in the urine of the rats. receiving high doses were found by H-1 NMR spectra. These may mean that high-dosage of ChangLe impairs the specific region of liver and kidney, such as renal tubule and mitochondria. The decrease in citrate and the increase in succinate and alpha-ketoglutarate were attributed to a combination of the inhibition of certain citric acid enzymes, renal tubular acidosis and the abnormal fatty acid catabolism. The information of the renal capillary necrosis could be derived from the increase in DMIA, DMG and TMAO. The increase in taurine was due to hepatic mitochondria dysfunction. The conclusions were supported by the results of biochemical measure. merits and enzymatic assay.

Species of rare earth in rat liver

The metabolic accumulation and species of rare earth in rat liver were investigated by ICP-MS and chromatography after the rats were fed by a low dose of mixed rare earth for a long time or the administration of a high dose of lanthanum for a short time. It was found that the content of rare earth in the liver increased with the arising of dose of drug delivery. Their accumulation rate was different, for example, La>Ce>Nd>Pr. The protein which could combine,with rare earth specially were not gotten through chromatography. It was suggested that rare earth could bind to many proteins voluntarily, such as some important enzymes and it might be separated from the combined proteins under certain conditions.

Species of lanthanum in Wistar rat liver

The metabolic accumulation and species of lanthanum in Wistar rat liver were investigated by ICP-MS, gel exclusion chromatography and ultrafiltration after the rats were fed by low dose of lanthanum for a long time. It was found that the content of La in the liver increased regularly with arise of dose and time of drug delivery. After the administration was stopped for a certain time a part of lanthanum in the liver Tvas metabolized, but;the metabolic rate was very slow, The lanthanum in rat liver was distributed in the soluble protein with molecular weight: of more than 60000 mostly. Rare Earth existed in the six elution peaks separated by Sephacryl S-200. The amount of lanthanum in the first elution fraction is the largest, which was 88 percent in the whole content of lanthanum in proteins with molecular weight more than 60000.

Effects of La3+ and Gd3+ on Ca2+ influx in rat hepatoma H-35 cells

Effects of La3+ and Gd3+ on Ca2+ influx were investigated in rat hepatoma H-35 cells by measuring the initial rate of Ca-45(2+) uptake. It was found that the maximum initial rate of Ca2+ uptake was increased six- to ten-fold at low concentrations of La3+ and Gd3+. Kinetic analyses by measuring the initial rate of Ca2+ influx at different external Ca2+ concentrations indicated the existence of two intracellular exchangeable components in the basal Ca2+ system, with low and high affinities for Ca2+, and only one class of Ca2+ binding sites was observed in the La3+- or Gd3+-treated cells. For high affinity, La3+ and Gc(3+) increased both kinetic parameters K-m and V-max of basal Ca2+ influx. La3+ and Gd3+ compete directly with Ca2+ for Ca2+ binding site for low affinity. The kinetics is competitive.

Dynamics of nonlinear error growth and season-dependent predictability of El Nino events in the Zebiak-Cane model

With the intermediate-complexity Zebiak-Cane model, we investigate the 'spring predictability barrier' (SPB) problem for El Nino events by tracing the evolution of conditional nonlinear optimal perturbation (CNOP), where CNOP is superimposed on the El Nino events and acts as the initial error with the biggest negative effect on the El Nino prediction. We show that the evolution of CNOP-type errors has obvious seasonal dependence and yields a significant SPB, with the most severe occurring in predictions made before the boreal spring in the growth phase of El Nino. The CNOP-type errors can be classified into two types: one possessing a sea-surface-temperature anomaly pattern with negative anomalies in the equatorial central-western Pacific, positive anomalies in the equatorial eastern Pacific, and a thermocline depth anomaly pattern with positive anomalies along the Equator, and another with patterns almost opposite to those of the former type. In predictions through the spring in the growth phase of El Nino, the initial error with the worst effect on the prediction tends to be the latter type of CNOP error, whereas in predictions through the spring in the decaying phase, the initial error with the biggest negative effect on the prediction is inclined to be the former type of CNOP error. Although the linear singular vector (LSV)-type errors also have patterns similar to the CNOP-type errors, they cover a more localized area than the CNOP-type errors and cause a much smaller prediction error, yielding a less significant SPB. Random errors in the initial conditions are also superimposed on El Nino events to investigate the SPB. We find that, whenever the predictions start, the random errors neither exhibit an obvious season-dependent evolution nor yield a large prediction error, and thus may not be responsible for the SPB phenomenon for El Nino events. These results suggest that the occurrence of the SPB is closely related to particular initial error patterns. The two kinds of CNOP-type error are most likely to cause a significant SPB. They have opposite signs and, consequently, opposite growth behaviours, a result which may demonstrate two dynamical mechanisms of error growth related to SPB: in one case, the errors grow in a manner similar to El Nino; in the other, the errors develop with a tendency opposite to El Nino. The two types of CNOP error may be most likely to provide the information regarding the 'sensitive area' of El Nino-Southern Oscillation (ENSO) predictions. If these types of initial error exist in realistic ENSO predictions and if a target method or a data assimilation approach can filter them, the ENSO forecast skill may be improved. Copyright (C) 2009 Royal Meteorological Society

Simultaneous determination of monoamine and amino acid neurotransmitters in rat endbrain tissues by pre-column derivatization with high-performance liquid chromatographic fluorescence detection and mass spectrometric identification

A sensitive and efficient method for simultaneous determination of glutamic acid (Glu), gamma-amino-butyric acid (GABA), dopamine (DA), 5-hydroxytryptamine (5-HT) and 5-hydroxyindole acetic acid (5-HIAA) in rat endbrains was developed by high-performance liquid chromatography (HPLC) with fluorescence detection and on-line mass spectrometric identification following derivatization with 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC). Different parameters which influenced derivatization and separation were optimized. The complete separation of five neurotransmitter (NT) derivatives was performed on a reversed-phase Hypersil BDS-C-18 column with a gradient elution. The rapid structure identification of five neurotransmitter derivatives was carried out by on-line mass spectrometry with electrospray ionization (ESI) source in positive ion mode, and the BCEOC-labeled derivatives were characterized by easy-to-interpret mass spectra. Stability of derivatives, repeatability, precision and accuracy were evaluated and the results were excellent for efficient HPLC analysis. The quantitative linear range of five neurotransmitters were 2.441-2 x 10(4) nM, and limits of detection were in the range of 0.398-1.258 nM (S/N = 3:1). The changes of their concentrations in endbrains of three rat groups were also studied using this HPLC fluorescence detection method. The results indicated that exhausting exercise could obviously influence the concentrations of neurotransmitters in rat endbrains. The established method exhibited excellent validity, high sensitivity and convenience, and provided a new technique for simultaneous analysis of monoamine and amino acid neurotransmitters in rat brain. (C) 2008 Elsevier B.V. All rights reserved.

LC determination of amino acids in rat plasma with fluorescence detection: Application to exercise physiology

An LC method for the determination of 20 amino acids (AAs), using 1,2-Benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC) as fluorescent labeling reagent, has been validated and applied for the analysis of AAs in rat plasma at three different states concerning exercise physiology. Identification of AA derivatives was carried out by LC-MS with electrospray ion (ESI), and the MS-MS cleavage mode of the representative tyrosine (Tyr) derivative was analyzed. Gradient elution on a Hypersil BDS C-18 column gave good separation of the derivatives. Excellent linear responses were observed and good compositional data could be obtained from as little as 50-200 mu L of plasma samples. The contents of 20 AAs in rat plasma of three groups (24 rats, group A: quiet state, group B: at exercising exhaust, group C: 12 h after exercising exhaust) exhibited evident difference corresponding to the physiological states. Facile BCEOC derivatization coupled with LC-FLD-ESI-MS analysis allowed the development of a highly sensitive method for the quantitative analysis of trace level of AAs from plasma or other biochemical samples.

Effects of hypoxia on contents of essential elements in pika and rat heart

The effects of hypoxia on the levels of essential macroelements and trace elements (K, Na, Ca, Mg, Cu, Zn, Fe, and Mn) in the heart muscles of Wistar rats and plateau pikas (Ochotona curzoniae) were studied by atomic absorption spectrometry. Unlike the rat, the plateau pika is tolerant to hypoxia. The levels of K, Na, and the trace element Mn were not significantly changed in rat or pika hearts after exposure to hypoxia for 1, 10, or 25 d at simulated altitudes of 5000 and 7000 m. Other minerals (Ca, Mg, Cu, Zn, and Fe) were significantly affected by hypoxia and the levels followed different time-courses under different hypoxic regimes in these two animals. There were marked differences between the rat and pika in myocardial accumulation of essential elements such as Ca, which was increased to high levels in the rat but not affected in the pika. The results suggest that hypoxia affects animal physiological mechanisms by regulating the levels of essential elements.