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General Physiology and Biophysics

Volume 32, 2013, No. 1


  Hydrogen sulfide in cell signaling, signal transduction, cellular bioenergetics and physiology in C. elegans
Katalin Módis 1), Katarzyna Wolanska, Roman Wozdek

1)Department of Anesthesiology, University of Texas Medical Branch, Galveston TX, USA.

Hydrogen sulfide (H2S), long viewed as a toxic gas and environmental hazard, is emerging as a biological mediator with remarkable physiological and pathophysiological relevance. H2S is now viewed as the third main gasotransmitter in the mammalian body. Its pharmacological characteristic possesses similarities to the other two gasotransmitters – nitric oxide (NO) and carbon monoxide (CO). Many of the biological effects of H2S follow a bell-shaped concentration-response; at low concentration or at lower release rates it has beneficial and cytoprotective effects, while at higher concentrations or fast release rates toxicity becomes apparent. Cellular bioenergetics is a prime example for this bell-shaped dose-response, where H2S, at lower concentrations/rates serves as an inorganic substrate and electron donor for mitochondrial ATP generation, while at high concentration it inhibits mitochondrial respiration by blocking the Complex IV in the mitochondrial electron transport chain. The current review is aimed to focus on the following aspects of H2S biology: 1) a general overview of the general pharmacological characteristics of H2S, 2) a summary of the key H2S-mediated signal transduction pathways, 3) an overview of role of H2S in regulation of cellular bioenergetics, 4) key aspects of H2S physiology in C. elegans (a model system) and, finally 5) the therapeutic potential of H2S donating molecules in various disease states.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 1-22.

  Dimethyl sulfoxide at high concentrations inhibits non-selective cation channels in human erythrocytes
Oleg Nardid 1), Miroslav Schetinskey, Yuliya Kucherenko

1)Institute for Problems of Cryobiology and Cryomedicine of the Ukrainian National Academy of Sciences, Kharkov 61015, Ukraine.

Dimethyl sulfoxide (DMSO), a by-product of the pulping industry, is widely used in biological research, cryobiology and medicine. On cellular level DMSO was shown to suppress NMDA-AMPA channels activation, blocks Na+ channel activation and attenuates Ca2+ influx (Lu and Mattson 2001). In the present study we explored the whole-cell patch-clamp to examine the acute effect of high concentrations of DMSO (0.1–2 mol/l) on cation channels activity in human erythrocytes. Acute application of DMSO (0.1–2 mol/l) dissolved in Cl–-containing saline buffer solution significantly inhibited cation conductance in human erythrocytes. Inhibition was concentration-dependent and had an exponential decay profile. DMSO (2 mol/l) induced cation inhibition in Cl–- containing saline solutions of: 40.3 ± 3.9% for K+, 35.4 ± 3.1% for Ca2+ and 47.4 ± 1.9% for NMDG+. Substitution of Cl– with gluconate– increased the inhibitory effect of DMSO on the Na+ current. Inhibitory effect of DMSO was neither due to high permeability of erythrocytes to DMSO nor to an increased tonicity of the bath media since no effect was observed in 2 mol/l glycerol solution. In conclusion, we have shown that high concentrations of DMSO inhibit the non-selective cation channels in human erythrocytes and thus protect the cells against Na+ and Ca2+ overload. Possible mechanisms of DMSO effect on cation conductance are discussed.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 23-32.

  Suppression of membrane vesiculation as anticoagulant and anti-metastatic mechanism. Role of stability of narrow necks
Roman Štukelj 1), Vid Šuštar, Anita Mrvar-Brečko, Peter Veranič, Henry Hägerstrand, Veronika Kralj-Iglič, France Sevšek

1)Laboratory of Clinical Biophysics, Faculty of Health Sciences, University of Ljubljana, Ljubljana, Slovenia.

Nanovesicles that are pinched off from biological membranes in the final stage of budding constitute a cell-cell communication system. Recent studies indicate that in vivo they are involved in blood clot formation and in cancer progression. The bud is connected to the mother membrane by a thin neck so it dwells close to the mother membrane. Using the electron microscopy we have observed in blood cells that adhesion between the membrane of the bud and of the mother cell in the vicinity of the neck took place and prevented the bud to pinch off from the mother vesicle. The same effect was observed in giant phospholipid vesicles (GPVs) due to attractive interaction between the bud and the mother vesicle mediated by the plasma protein beta-2-glycoprotein I. The stability of the neck is important for this process. By using Fourier method we analyzed thermal fluctuations of a GPV while a protrusion composed of beads connected by thin necks was spontaneously integrated into the mother GPV. Stepwise change of Fourier coefficients indicates an increased stability of necks which contributes to the retention of buds by the mother membrane and promotes anticoagulant and anti-metastatic mechanism by suppression of nanovesiculation.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 33-45.

  Effects of selenoprotein P on the contraction and relaxation of the airway smooth muscle
Sachie Ogawa 1), Min-Chul Shin, Masaki Hirashima, Norio Akaike, Yushi Ito

1)Research Division for Life Sciences, Kumamoto Health Science University, 325 Izumi-machi, Kumamoto 861-5598, Japan.

Selenoprotein P (SeP) not only represents the major selenoprotein in plasma, but also provides more than 50% of the total plasma selenium. However, there is no report concerning the direct action of selenium or selenium-containing compounds on the contraction and relaxation of the airway smooth muscle. Therefore, we investigated the effects of SeP and sodium selenite (SS) on the indirectly induced contraction and relaxation of the cat bronchi, and gel contraction of cultured bovine tracheal smooth muscle cells (BTSMC) induced by ATP. In the present results, SeP or SS suppressed the amplitude of twitch-like contractions of cat bronchiole without affecting the non-adrenergic and non-cholinergic (NANC) relaxations evoked by electrical field stimulation. SeP also suppressed the ATP-induced gel contraction of BTSMC. These results suggest that SeP suppresses the amplitude of twitch-like contraction of cat bronchiole by acting directly on the bronchiolar smooth muscle.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 47-54.

  Analysis of Leucine-rich repeat kinase 2 (LRRK2) and Parkinson protein 2 (parkin, PARK2) genes mutations in Slovak Parkinson disease patients
Csaba Bognar 1), Marian Baldovic, Jan Benetin, Ludovit Kadasi, Andrea Zatkova

1)Department of Molecular Biology, Faculty of Natural Sciences, Comenius University, Bratislava, Slovak Republic.

Parkinson disease (PD) is a chronic neurodegenerative movement disorder characterized by selective loss of nigrostriatal dopaminergic neurons and formation of Lewy bodies. Clinical manifestations include motor impairments involving tremor, bradykinesia, postural instability and rigidity. Using dHPLC method we screened exons 31, 35, 41, 48 of the Leucine-rich repeat kinase 2 (LRRK2) gene and exons 2, 6 and 7 of Parkinson protein 2 (parkin, PARK2) genes in a cohort of 216 consecutive, unrelated Slovak patients with familial or sporadic PD, including early and late onset. By this means we aimed to detect the most common pathogenic mutations within LRRK2 (Arg1441Cys, Arg1441Gly, Arg1628Pro, Tyr1699Cys, Gly2019Ser, Ile2020Thr, Gly2385Arg) and parkin genes responsible for late and early onset forms of disease, respectively. However, none of these mutations was identified in our cohort. Heterozygous point mutation p.Arg275Trp in exon 7 of parkin gene was identified in one patient with age at onset 61 years. Furthermore, we observed the presence of one exonic (LRRK2 ex 48: 7155A>G) and eight intronic polymorphisms (in LRRK2: IVS35+23T>A, IVS47-91insGCCAT, IVS47-91insGCAT, IVS47-41A>G, IVS47-9delT, IVS47-20C>T, IVS47-90A>G, in parkin: IVS2+25T>C), three of which were novel.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 55-66.

  A viscometric approach of pH effect on hydrodynamic properties of human serum albumin in the normal form
Karol Monkos 1)

1)Department of Biophysics, Medical University of Silesia, H. Jordana 19, 41-808 Zabrze 8, Poland.

The paper presents the results of viscosity determinations on aqueous solutions of human serum albumin (HSA) at isoelectric point over a wide range of concentrations and at temperatures ranging from 5°C to 45°C. On the basis of a modified Arrhenius equation and Mooney’s formula some hydrodynamic parameters were obtained. They are compared with those previously obtained for HSA in solutions at neutral pH. The activation energy and entropy of viscous flow and the intrinsic viscosity reach a maximum value, and the effective specific volume, the self-crowding factor and the Huggins coefficient a minimum value in solutions at isoelectric point. Using the dimensionless parameter [η]c, the existence of three ranges of concentrations: diluted, semi-diluted and concentrated, was shown. By applying Lefebvre’s relation for the relative viscosity in the semi-dilute regime, the Mark-Houvink-Kuhn-Sakurada (MHKS) exponent was established. The analysis of the results obtained from the three ranges of concentrations showed that both conformation and stiffness of HSA molecules in solutions at isoelectric point and at neutral pH are the same.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 67-78.

  Melatonin protects rat thymus against oxidative stress caused by exposure to microwaves and modulates proliferation/apoptosis of thymocytes
Dusan Sokolovic 1), Branka Djordjevic, Gordana Kocic, Andrej Veljkovic, Milena Marinkovic, Jelena Basic, Tatjana Jevtovic-Stoimenov, Zoran Stanojkovic, Danka Sokolovic, Voja Pavlovic, Boris Djindjic, Dejan Krstic

1)Department of Biochemistry, Faculty of Medicine, University of Nis, bul. Dr. Zorana Djindjica 81, 18000 Nis, Serbia.

The aim of the study was to evaluate the effect of melatonin on oxidative stress, DNA fragmentation, apoptsis and proliferation in thymus tissue of rats exposed to microwaves. Wistar rats were divided in four groups: I – treated with saline; II – treated with melatonin; III – microwaves exposed; IV – microwaves exposed and melatonin treated. Melatonin (2 mg/kg i.p.) was administered daily. Animals were sacrificed after 20, 40 and 60 days. A significant increase in malondialdehyde and carbonyl group content, as well as decrease in catalase and increase in xanthine oxidase activity were registered under microwave exposure. Melatonin prevented the increase in malondialdehyde and carbonyl group content, and reversed the effect on catalase and xanthine oxidase activity. Both, alkaline and acid DNase activity were increased due to microwave exposure. Furthermore, microwaves caused increase in apoptosis rate (detected using Annexin V-FITC/PI kit) and reduced proliferative capacity of thymocytes (induced by ConA). However, melatonin caused decrease in alkaline and acid DNase activity, decrease in apoptotic rate and increase in proliferation rate of thymocytes. Melatonin exerts protective effects on rat thymocytes by modulating processes of apoptosis and proliferation, and causes decrease in DNA fragmentation and oxidative stress intensity under exposure to microwaves.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 79-90.

  Long-term alterations of cell population in the adult rat forebrain following the exposure to fractionated doses of ionizing radiation
Soňa Bálentová 1), Eva Hajtmanová, Ivana Kinclová, Ján Lehotský, Dušan Dobrota, Marián Adamkov

1)Institute of Histology and Embryology, Jessenius Faculty of Medicine, Comenius University, Malá Hora 4, 036 01 Martin, Slovak Republic.

We investigated radiation-induced delayed alterations of proliferating population, cells undergoing apoptosis and glial cells housed rat brain neurogenic region. Adult male Wistar rats were investigated 30, 60 or 90 days after whole-body irradiation with fractionated doses of gamma rays (the total dose of 4 Gy). Using immunohistochemistry for detection of cell proliferation marker Ki-67, caspase3 as apoptotic marker and GFAP for mature astrocytes we have been performed quantitative analysis in different forebrain's areas along the SVZ-OB axis, i.e. in the anterior subvetricular zone (SVZa), vertical arm, elbow and horizontal arm. In animals that survived thirty days after radiation treatment initial decrease of the Ki-67-positive cells was seen in regions along the SVZ-OB axis. The highest increase was observed in vertical arm on the 60th day followed by the most striking decline on the 90th day after irradiation. Cells undergoing apoptosis didn't showed expressive increase during entire experiment except of horizontal arm. The most striking changes of GFAP-positive cells were seen 30 and 60 days after irradiation in vertical arm and elbow. Results suggested that radiation response of proliferating cells and astrocytes resides the SVZa may play contributory role in development of more adverse radiation-induced late effects.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 91-100.

  GABA-induced vasorelaxation mediated by nitric oxide and GABAA receptor in non diabetic and streptozotocin-induced diabetic rat vessels
Mahmoud Kamran 1), Abdolreza Bahrami, Nepton Soltani, Mansoor Keshavarz, Leila Farsi

1)Physiology Department, Faculty of Medicine, and Research Center for Molecular Medicine, Hormozgan University of Medical Science, Bandar Abbas, Iran.

Diabetes has profound, negative effects on the function of arteries and arterioles. Hypertension is considered an independent risk factor for cardiovascular mortality in diabetic patients. The present study was designed to determine whether GABA-induced vasorelaxation in normal and streptozotocin-induced diabetic rat vessels is mediated by nitric oxide and the GABAA receptor. Diabetes was induced by a single intraperitoneal injection of streptozotocin (STZ) (60 mg/kg). Eight weeks later, superior mesenteric arteries of all groups were isolated and perfused according to the McGregor method. Baseline perfusion pressure of STZ diabetic rats was significantly higher than non-diabetic rats in both intact and denuded endothelium. In the presence of bicuculline, a selective GABAA receptor blocker, GABA-induced relaxation in intact and denuded endothelium mesenteric beds of non-diabetic and STZ diabetic rats was suppressed. But in the presence of L-NAME, a nitric oxide synthesis inhibitor, although GABA- induced vasorelaxation was not suppressed at a dose of 1 µM of GABA in all groups, this response was suppressed with the other doses of GABA. From the results of this study it may be concluded that the vasorelaxatory effect of GABA is mediated by the GABAA receptor and nitric oxide in both STZ diabetic and non-diabetic vessels.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 101-106.

  5-HT1A receptor-mediated activation of outward potassium current by serotonin in mouse cultured spiral ganglion neurons
Chao Yu 1), Jin Liu, Yanfang Chen, Yuedi Tang

1)Department of Otorhinolaryngology and Translational Neuroscience Center, West China Hospital of Sichuan University, No. 37 Guo-Xue-Xiang, Chengdu, Sichuan 610041, P. R. China.

Cochlear spiral ganglion neurons provide the only pathway for transmitting sound evoked activity from the hair cells to the central auditory system. Serotonin plays a role in the response properties of central auditory neurons. However, knowledge about the role of serotonin in the peripheral auditory nervous system remains limited. In the current study, we investigated the influence of serotonin on outward potassium current in mouse cultured spiral ganglion neurons using whole-cell patch clamp technique. The cell capacitance was 4.03 ± 0.18 pF (n = 54). Application of serotonin caused an increase of outward potassium currents within seconds, whereas treatment with WAY100635, a selective 5-HT1A receptor antagonist, counteracted the increase effect of serotonin. These results suggest that serotonin increases outward potassium currents in cultured spiral ganglion neurons through the activation of 5-HT1A receptor. Serotonin may play an important role in sound transmission.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 107-114.

  Phytochemical, antioxidant and protective effect of Rhus tripartitum root bark extract against ethanol-induced ulcer in rats
Hichem Alimi 1), Sakhria Mbarki, Zeineb Barka, Anwer Feriani, Zouhour Bouoni, Najla Hfaeidh, Mohsen Sakly, Olfa Tebourbi, Khémais Rhouma

1)Research unit of Macromolecular Biochemistry and Genetic, Faculty of Sciences of Gafsa, 2112 Gafsa, Tunisia.

Rhus tripartitum (sumac) is an Anacardiaceae tree with a wide phytotherapeutic application including the use of its roots in the management of gastric ulcer. In the present study the Rhus tripartitum root barks extract (RTE) was phytochemical studied, in vitro tested for their potential antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and reducing power assay and in vivo evaluated for its ability to prevent ethanol-induced gastric ulcer in rats. The RTE was rich in phenolics, flavonoids, tannins and polysaccharide contents and exhibited a low but not weak in vitro antioxidant activity when compared with (+)-catechin. Pre-treatment with RTE at oral doses 50, 200 and 400 mg/kg body weight was found to provide a dose-dependent protection against ethanol-induced ulcer by averting the deep ulcer lesions of the gastric epithelium, by reducing gastric juice and acid output, by enhancing gastric mucus production by preserving normal antioxidant enzymes activities, and inhibiting the lipid peroxidation. The antiulcerogenic activity of RTE might be due to a possible synergistic antioxidant and antisecretory effects.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 115-127.

  Maternal restraint stress negatively influences growth capacity of preimplantation mouse embryos
Ján Burkuš 1), Štefan Čikoš, Dušan Fabian, Janka Kubandová, Soňa Czikková, Juraj Koppel

1)Institute of Animal Physiology, Slovak Academy of Sciences, Šoltésovej 4, 040 01 Košice, Slovak Republic.

In our study we investigated the effect of maternal restraint stress on preimplantation embryo development using a mouse model. We exposed hormonally stimulated (superovulated) and unstimulated (i.e. spontaneously ovulating) mouse females to restraint stress for 30 min three times a day during the preimplantation period. The stress exposure caused significant increase in blood plasma corticosterone concentration. Microscopical evaluation of embryos isolated from spontaneously ovulating females showed that maternal stress significantly increased the proportion of embryos with lower cell numbers (≤32 cells) and decreased the proportion of embryos with higher cell numbers (65–96 cells and 97–128 cells). Moreover maternal restraint stress decreased the cell counts per embryo and per blastocyst. After an additional 24 h in vitro culture we did not find any difference in the embryo distribution or in the cell counts per embryo/blastocyst between embryos isolated from stressed and control mothers. The exposure to restraint stress did not affect the incidence of apoptosis in blastocysts isolated from spontaneously ovulated dams. In gonadotropin stimulated dams, the hormonal treatment itself notably changed embryo distribution (increasing the proportion of degenerated embryos) and increased the occurrence of apoptotic cells. Our results indicate that psychical stress exposure in very early pregnancy can significantly influence the developmental capacity of preimplantation embryos.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 129-137.

  Brain-derived neurotrophic factor blood levels in two models of transient brain ischemia in rats
Miroslav Gottlieb 1), Petra Bonova, Viera Danielisova, Miroslava Nemethova, Jozef Burda , Dasa Cizkova

1)Institute of Neurobiology, Slovak Academy of Sciences, Kosice, Slovak Republic.

We monitored possible influence of transient focal and global brain ischemia on BDNF blood level. In both models noticeable fluctuation of BDNF concentration mainly in reperfusion was observed. During the first 90 min, BDNF in total blood and in blood cells continuously decreased in both models but plasma BDNF raised at 40 min and peaked at 90 min of reperfusion. Our data confirm the impact of transient brain ischemia on BDNF levels in the circulatory system, suggest blood cells as a possible source of BDNF and demonstrate the interdependence of blood compartments and physiological state of an affected organism.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 139-142.

  Excluded volume effect of counterions and water dipoles near a highly charged surface due to a rotationally averaged Boltzmann factor for water dipoles
Ekaterina Gongadze 1), Aleš Iglič

1)Laboratory of Biophysics, Faculty of Electrical Engineering, University of Ljubljana, Tržaška cesta 25, Ljubljana, Slovenia.

Water ordering near a negatively charged electrode is one of the decisive factors determining the interactions of an electrode with the surrounding electrolyte solution or tissue. In this work, the generalized Langevin-Bikerman model (Gongadze-Iglič model) taking into account the cavity field and the excluded volume principle is used to calculate the space dependency of ions and water number densities in the vicinity of a highly charged surface. It is shown that for high enough surface charged densities the usual trend of increasing counterion number density towards the charged surface may be completely reversed, i.e. the drop in the counterions number density near the charged surface is predicted.

General Physiology and Biophysics. Volume 32, 2013, No. 1: 143-145.