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General Physiology and Biophysics


Volume 30, 2011, No. 1

Content:


  B cells – ontogenesis and immune memory development
Zuzana Nescakova 1), Slavomir Bystricky

1)Institute of Chemistry, Glycomed Centre, Slovak Academy of Sciences, Dúbravská cesta 9, 845 38 Bratislava, Slovakia. chemnesc@savba.sk.


Functional diversity in the distinct developmental stages as well as anti-pathogen effectiveness and memory functionality make B cells unique and attractive object of physiology studies of the immune system. B cells are produced throughout the life of an organism, originate from the hematopoietic stem cells before birth and continue to differentiate to terminal stages. Over the past decade, there has been considerable progress in the research of all B cell intermediates and developmental processes. In this review, we will try to bring brief and comprehensive description of the current understanding of this fascinating topic.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 1-10.

 
  Effects of aldosterone and mineralocorticoid receptor antagonism on cardiac ion channels in the view of upstream therapy of atrial fibrillation
Roman Laszlo 1), Kerstin Bentz, Juergen Schreieck

1)Medizinische Klinik III, Kardiologie und Kreislauferkrankungen, Eberhard Karls Universität Tuebingen, Otfried-Mueller-Strasse 10, D 72076 Tuebingen, Germany. roman.laszlo@gmx.de.


Atrial fibrillation (AF) is the most common sustained arrhythmia in man. Over the past years, importance of the renin-angiotensin-aldosterone system in AF pathophysiology has been recognized. Lately, the role of aldosterone in AF pathophysiology and mineralocorticoid receptor (MR) antagonism in “upstream” AF treatment is discussed with special regards concerning the effects on AF-induced structural remodeling. However, there is more and more evidence that MR antagonism also influences atrial electrophysiology and, respectively, AF-induced electrical remodeling, whereas the molecular mechanisms are almost unknown. The aim of this mini-review is to give an overview about the role of aldosterone in AF pathophysiology in principle and to summarize current available data concerning affection of cardiac ion channels by aldosterone and MR antagonism. Finally, as modulation of oxidative stress is discussed as one main therapy principle of “upstream” treatment of AF, potential mechanisms how modulation of oxidative stress by aldosterone and accordingly MR antagonism might alter atrial ion currents are delineated. Summarized, publications concerning potential mechanisms of aldosterone- and MR antagonism-modulated cardiac ion channels in various experimental settings are almost exclusively limited to the ventricular level and, partly, they are also contradictorily. Translation of these data to the atria is problematic because atrial and ventricular electrophysiology exhibit remarkable differences. It can be concluded that further research on the “atrial level” is needed in order to clarify the potential impact of the affection of atrial ion channels by aldosterone and accordingly MR antagonism in “upstream” therapy of AF.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 11-19.

 
  The role of BKCa channels on hyperpolarization mediated by hyperosmolarity in human articular chondrocytes
Julio Sánchez 1), Diego López-Zapata

1)Laboratorio de Fisiología Celular, Departamento de Ciencias Básicas, Facultad Ciencias de la Salud, Universidad Tecnológica de Pereira, La Julita, Pereira AA 97, Colombia. jcsanchez@utp.edu.co.


Chondrocytes, the only cell in cartilage, are subjected to hyperosmotic challenges continuously since extracellular osmolarity in articular cartilage increases in response to mechanical loads during joint movement. Hyperosmolarity can affect membrane transport, and it is possible that load modulates matrix synthesis through alterations in intracellular composition. In the present study, the effects of hyperosmotic challenges were evaluated using the whole-cell patch clamp technique, whole cell mode on freshly isolated human and bovine articular chondrocytes. In human chondrocytes, hypertonicity induced the activation of outward Ca2+-sensitive K+ currents, which were inhibited by iberiotoxin and TEA-Cl. The current induced by hypertonic switching (osmolarity from 300 to 400 mOsm/l) caused cell hyperpolarization (from –39 mV to –70 mV) with a reversal potential of –96 ± 7 mV. These results suggest a role for Ca2+-activated K+ channels in human articular chondrocytes, leading to hyperpolarization as a consequence of K+ efflux through these channels. These channels could have a role in the articular chondrocyte’s response to a hyperosmotic challenge and matrix metabolism regulation by load.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 20-27.

 
  Contribution of Nav1.1 to cellular synchronization and automaticity in spontaneous beating cultured neonatal rat ventricular cells
Haruko Masumiya 1)

1)Division of Physiome, Department of Physiology, Hyogo College of Medicine, Nishinomiya, Hyogo, 663-8501 Japan. masumiya@hyo-med.ac.jp.


Two factors of cell coupling influence cellular synchronization and automaticity: gap junction coupling and ion channels activity. However, the role of Na+ channel isoforms underlying cell-to-cell interaction and cellular automaticity is not well understood. To address these questions, we studied mRNA expression of Na+ channel isoforms and the effects of TTX on spontaneously beating cultured ventricle cells. Using RT-PCR technique we demonstrated the presence of Nav1.1 and Nav1.5 channels. The reduction of Nav1.1 channel activity disturbed cell-to-cell interaction and changed beating rates. Thus, Nav1.1 channel is involved in cellular synchronization and automaticity.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 28-33.

 
  Role of glucagon-like peptide-1 and its agonists on early prevention of cardiac remodeling in type 1 diabetic rat hearts
Ghinwa Barakat 1), Nuha Nuwayri-Salti, Lina Kadi, Khalil Bitar, Wael Al-Jaroudi, Anwar Bikhazi

1)Department of Physiology, Diana Tamari Sabbagh/2-44, American University of Beirut, Medical Center, P.O.Box 11-0236, Riad El Solh, Beirut 1107 2020, Lebanon. anwar.bikhazi@aub.edu.lb.


Glucagon-like peptide-1 (GLP-1) is an incretin hormone secreted from intestinal L cells upon nutrients ingestion, and is currently used for treating diabetes mellitus. It plays an important role in receptor modulation and cross talk with insulin at the coronary endothelium (CE) and cardiomyocytes (CM) in diabetic type 1 rat heart model. We studied the effects of insulin, GLP-1 analogues (exendin-4), and dipeptidyl peptidase-IV (DPP-IV) inhibitor on GLP-1 cardiac receptor modulation. The binding affinity of GLP-1 to its receptor on CE and CM was calculated using a rat heart perfusion model with [125I]-GLP-1(7-36). Tissue samples from the heart were used for immunostaining and Western blot analyses. GLP-1 systemic blood levels were measured using ELISA. GLP-1 binding affinity (τ) increased on the CE (0.33 ± 0.01 vs. 0.25 ± 0.01 min; p < 0.001) and decreased on the CM (0.29 ± 0.02 vs. 0.43 ± 0.02 min; p < 0.001) in the diabetic non-treated rats when compared to normal. There was normalization of τ back to baseline on the CE and CM levels with insulin and DPP-IV inhibitor treatment, respectively. Histological sections and immunofluorescence showed receptor up-regulation in diabetic rats with significant decrease and even normalization with the different treatment strategies. Systemic GLP-1 levels increased after 14 days of diabetes induction (10 ± 3.7 vs. 103 ± 58 pM; p = 0.0005). In conclusion, there is a significant GLP-1 receptor affinity modulation on the CE and CM levels in rats with diabetes type 1, and a cross talk with GLP-1 analogues in early prevention of cardiac remodeling.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 34-44.

 
  Specificity of the volume-activated amino acid efflux pathway in cultured human breast cancer cells
David Shennan 1), Jean Thomson

1)Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Royal College, 204 George Street, Glasgow, G1 1XW, United Kingdom. Dtaurine@aol.com.


It has been shown that cell swelling stimulates the efflux of taurine from MCF-7 and MDA-MB-231 cells via a pathway which has channel-like properties. The purpose of this study was to examine the specificity of the volume-activated taurine efflux pathway in both cell lines. A hyposmotic shock increased the efflux of glycine, L-alanine, AIB (α-aminoisobutyric acid), D-aspartate but not L-leucine from MDA-MB-231 and MCF-7 cells. It was evident that the time course of activation/inactivation of those amino acids whose efflux was affected by cell swelling was similar to that of volume-activated taurine efflux. The effect of exogenous ATP on swelling-induced glycine, AIB and D-aspartate efflux from MDA-MB-231 cells was similar to that found on taurine efflux. In addition, volume-activated AIB efflux from MDA-MB-231 cells, like that of swelling-induced taurine efflux, was inhibited by diiodosalicylate. Tamoxifen inhibited volume-activated taurine release from both MDA-MB-231 and MCF-7 cells. The results suggest that neutral and anionic α-amino acids are able to utilize the volume-activated taurine efflux pathway in both cell lines. The effect of tamoxifen on breast cancer growth may, in part, be related to perturbations in cell volume regulation.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 45-51.

 
  Effect of cadmium and lead on the membrane potential and photoelectric reaction of Nitellopsis obtusa cells
Renata Kurtyka 1), Zbigniew Burdach, Waldemar Karcz

1)Department of Plant Physiology, Faculty of Biology and Environmental Protection, University of Silesia, ul. Jagiellońska 28, 40 032 Katowice, Poland. renata.kurtyka@us.edu.pl.


The effects of Cd and Pb on membrane potential (Em) and photoelectric reaction of Nitellopsis obtusa cells were investigated. It was found that Cd and Pb at 1.0 mM caused a depolarization of the Em, whereas both metals at lower concentrations changed the Em in a different way. Pb at 0.1 mM and 0.01 mM hyperpolarized the Em, whereas Cd at the same concentrations depolarized and did not change the Em, respectively. In the presence of 0.01 mM Pb, the light-induced hyperpolarization of the Em was by 18% higher as compared to the control, whereas at 1.0 mM Pb it was by 40% lower. Pb at 0.1 mM and Cd at 0.01 mM or 5 × 0.01 mM did not change the light-induced membrane hyperpolarization. However, in the presence of Cd at 0.1 mM and 1.0 mM this hyperpolarization was 2-fold lower or was completely abolished, respectively. These results suggest that at high Cd and Pb concentrations both depolarization of the Em and decrease of light-induced membrane hyperpolarization in Nitellopsis obtusa cells are probably due to inhibition of the plasma membrane H+-ATPase activity, whereas both metals at lower concentrations differ in mechanism of membrane potential changes.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 52-58.

 
  Transient finite element modeling of functional electrical stimulation
Nenad Filipovic 1), Aleksandar Peulic, Nebojsa Zdravkovic, Vesna Grbovic-Markovic, Aleksandra Jurisic-Skevin

1)Faculty of Mechanical Engineering, University of Kragujevac, S. Janjic 6, 34 000 Kragujevac, Serbia. fica@kg.ac.rs.


Transcutaneous functional electrical stimulation is commonly used for strengthening muscle. However, transient effects during stimulation are not yet well explored. The effect of an amplitude change of the stimulation can be described by static model, but there is no differency for different pulse duration. The aim of this study is to present the finite element (FE) model of a transient electrical stimulation on the forearm. Discrete FE equations were derived by using a standard Galerkin procedure. Different tissue conductive and dielectric properties are fitted using least square method and trial and error analysis from experimental measurement. This study showed that FE modeling of electrical stimulation can give the spatial-temporal distribution of applied current in the forearm. Three different cases were modeled with the same geometry but with different input of the current pulse, in order to fit the tissue properties by using transient FE analysis. All three cases were compared with experimental measurements of intramuscular voltage on one volunteer.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 59-65.

 
  Deteriorating effect of fluvastatin on the cholestatic liver injury induced by bile duct ligation in rats
Halka Lotková 1), Pavla Staňková, Tomáš Roušar, Otto Kučera, Lukáš Kohoutek, Stanislav Mičuda, Eva Brčáková, Gabriela Kolouchová, Zuzana Červinková

1)Department of Physiology, Faculty of Medicine in Hradec Králové, Charles University in Prague, Šimkova 870, 500 38 Hradec Králové, Czech Republic. lotko@lfhk.cuni.cz.


Antiinflammatory effect of statins mediated by the reduction of cytokine IL-6 in hepatocytes have been reported. Contrary to beneficial effect, statins can increase susceptibility to mitochondrial dysfunction. Extrahepatic biliary obstruction is associated with oxidative stress, pro-inflammatory response and hepatocyte mitochondrial dysfunction. The aim of our study was to verify the effect of fluvastatin on cholestatic liver injury. Cholestasis was induced in Wistar rats by bile duct ligation. Fluvastatin (1 or 5 mg/kg) was administered after surgery and then daily for 7 days. The dose of 5 mg/kg led to the deterioration of hepatocellular injury. Despite lower production of IL-6, decrease in GSH content, rise of TGFß and inhibition of respiratory complex I in mitochondria were determined. The mRNA expressions of canalicular transporter Mdr1b and basolateral transporter Mrp3 increased in cholestatic liver. Fluvastatin administration then led to the attenuation of this change. Analogously, mRNA expression of conjugative enzyme Ugt1a1 was diminished by fluvastatin administration to cholestatic rats. We can conclude that decrease in the antioxidative status and mitochondrial dysfunction could at least in part participate on the deteriorating effect of fluvastatin. Whether these processes can be a consequence of the alteration in metabolism and transport of potentially toxic substances remains to verify.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 66-74.

 
  Competition of NO synthases and arginase in the airways hyperreactivity
Anna Strapkova 1), Martina Antošová

1)Department of Pharmacology, Jessenius Faculty of Medicine, Comenius University, Sklabinská 26, 037 53 Martin, Slovakia. astrapkova@jfmed.uniba.sk.


The competition between arginases and NO synthases (NOS) for their common substrate L-arginine can be important in the airways hyperreactivity. We investigated the effect of the simultaneous modulation of arginase and NOS activities in allergen-induced airways hyperreactivity. We analysed the response of tracheal and lung tissue smooth muscle to histamine or acetylcholine after administration Nω-nitro-L-arginine methyl ester (L-NAME), aminoguanidine (AG) and Nω-hydroxy-L-arginine (NOHA) in the combinations in in vitro conditions. The results show the decrease of ovalbumin-induced hyperreactivity after inhibition of arginase activity with NOHA. A supplementation of L-arginine caused favourable effect on the airway smooth muscle response. We found the airway reactivity decrease on the whole if we used the combination of NOS and arginase inhibitors. The inhibition of both types of enzymes caused more expressive effect in tracheal smooth muscles. We recorded the difference in the response to histamine or acetylcholine. The simultaneous inhibition of iNOS (with AG) and arginase (with NOHA) evoked the most expressive effect. Results show the importance of competition of both types enzymes – NOS and arginase for the balance of theirs activities in the control of airways bronchomotoric tone in the conditions of the airways hyperreactivity.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 75-83.

 
  900 MHz pulse-modulated radiofrequency radiation induces oxidative stress on heart, lung, testis and liver tissues
Meric Esmekaya 1), Cigdem Ozer, Nesrin Seyhan

1)Department of Biophysics, Gazi University, Faculty of Medicine, 06510 Beşevler, Ankara, Turkey. mericarda@yahoo.com.


Oxidative stress may affect many cellular and physiological processes including gene expression, cell growth, and cell death. In the recent study, we aimed to investigate whether 900 MHz pulse-modulated radiofrequency (RF) fields induce oxidative damage on lung, heart and liver tissues. We assessed oxidative damage by investigating lipid peroxidation (malondialdehyde, MDA), nitric oxide (NOx) and glutathione (GSH) levels which are the indicators of tissue toxicity. A total of 30 male Wistar albino rats were used in this study. Rats were divided randomly into three groups; control group (n = 10), sham group (device off, n = 10) and 900 MHz pulsed-modulated RF radiation group (n = 10). The RF rats were exposed to 900 MHz pulsed modulated RF radiation at a specific absorption rate (SAR) level of 1.20 W/kg 20 min/day for three weeks. MDA and NOx levels were increased significantly in liver, lung, testis and heart tissues of the exposed group compared to sham and control groups (p < 0.05). Conversely GSH levels were significantly lower in exposed rat tissues (p < 0.05). No significantly difference was observed between sham and control groups. Results of our study showed that pulse-modulated RF radiation causes oxidative injury in liver, lung, testis and heart tissues mediated by lipid peroxidation, increased level of NOx and suppression of antioxidant defense mechanism.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 84-89.

 
  Beta-3 agonist-induced lipolysis and nitric oxide production: relationship to PPARgamma agonist/antagonist and AMP kinase modulation
Jiří Hodis 1), Radka Vaclavíková, Hassan Farghali

1)Institute of Pharmacology, First Faculty of Medicine, Charles University, Albertov 4, 120 00 Prague 2, Czech Republic. hodis@atlas.cz.


PPARgamma receptor agonist –troglitazone increases insulin sensitivity in visceral adipocytes and also increases fat mass. Beta-3 adrenergic receptor agonists mediate lipolysis and NO production (iNOS transcription) in visceral adipocytes. Troglitazone could possibly interfere with Beta-3-triggered lipolysis. We tested the crosstalk between PPARgamma agonist and Beta-3 agonist pathways on lipolysis and NO production in first 24 hours of treatment. Isolated epididymal rat adipocytes were cultivated in DMEM for 24 hours with treatment with Beta-3 agonist – BRL-37344, PPARgamma agonist – troglitazone, PPARgamma antagonist – SR-202 and AMPK blocker – compound C alone as well as in combinations. After 24 hours, lipolysis was measured by free glycerol, NO production by Griess reagent and iNOS mRNA by qRT-PCR. BRL-37344 increased lipolysis and NO production with iNOS transcription. Troglitazone increased all the three parameters as well but less than BRL-37344. Combination of troglitazone or SR-202 with BRL-37344 decreased NO production, iNOS transcription and lipolysis triggered before adding of BRL-37344. Compound C completely blocked the effect of troglitazone (and SR-202 as well) on BRL-37344. PPARgamma agonist/antagonist interferes with Beta-3 agonist activity in 24 hours. Troglitazone/SR-202 effect on Beta-3 triggered lipolysis and iNOS mRNA production is probably not PPAR gamma- but rather AMPK-dependent in first 24 hours (AMPK blocker – compound C blocked the effect).

General Physiology and Biophysics. Volume 30, 2011, No. 1: 90-99.

 
  Pharmacological studies of Cav3.1 T-type calcium channels using automated patch-clamp techniques
Kee Choi 1), Chiman Song, Chan Cheong, Hyewhon Rhim

1)Life/Health Division, Korea Institute of Science and Technology, 39-1 Hawolgok-Dong, Seongbuk-Gu, Seoul 136-791, Korea. keehyun@kist.re.kr.


T-type calcium channels are involved in a variety of physiological and pathophysiological processes, and thus could be therapeutic targets. However, there is no T-type channel selective blocker for use in clinical practice, demanding a need for the development of novel drugs where a higher-throughput screening system is required. Here we present pharmacological studies on Cav3.1 T-type channels using automated patch-clamp. The IC50 values obtained from automated patch-clamp and conventional one showed a good correlation (correlation coefficient of 0.82), suggesting that the automated patch-clamp is an efficient and reliable method for ranking the drug potencies for T-type channels.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 100-105.

 
  Evaluation of uridine 5’-eicosylphosphate as a stimulant of cyclic AMP-dependent cellular function
Masahiro Yutani 1), Akira Ogita, Ken-ichi Fujita, Yoshinosuke Usuki, Toshio Tanaka

1)Graduate School of Science, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585, Japan. tanakato@sci.osaka-cu.ac.jp.


Sporulation of the yeast Saccharomyces cerevisiae is negatively regulated by cyclic AMP (cAMP). This microbial cell differentiation process was applied for the screening of a substance that can elevate the intracellular cAMP level. Among nucleoside 5’-alkylphosphates, uridine 5’-eicosylphosphate (UMPC20) selectively and predominantly inhibited ascospore formation of the yeast cells. We suppose the inhibitory effect of UMPC20 could indeed reflect the elevation of the cellular cAMP level.

General Physiology and Biophysics. Volume 30, 2011, No. 1: 106-109.