T Márián

Publications of T Márián

• A PET study on the characterization of partially reversible radiogenic lower motor neurone disease.

  Authors: O Esik, Z Lengyel, G Sáfrány, K Vönöczky, P Agoston, J Székely, E Lengyel, T Márián, L Trón, I Bodrogi

  Spinal cord : the official journal of the International Medical Society of Paraplegia. 10/2002; 40(9):468-73.

  OBJECTIVE: To investigate the pathomechanism of the rare radiogenic lower motor neurone disease (LMND) on the basis of a case history involving a partial functional recovery. PATIENT: A 31-year-oldOBJECTIVE: To investigate the pathomechanism of the rare radiogenic lower motor neurone disease (LMND) on the basis of a case history involving a partial functional recovery. PATIENT: A 31-year-old seminoma patient received postoperative para-aortic and para-iliac telecobalt irradiation with a biologically effective dose of 88 Gy(2) (44 Gy in 2 Gy fractions/day, with an estimated alpha/beta of 2 Gy) delivered to the spinal cord following a single cycle of chemotherapy. LMND developed 4 months after the completion of radiotherapy. The patient exhibited flaccid paraparesis of the lower extremities (without sensory or vegetative signs), followed by a worsening after further chemotherapy, due to pulmonary metastatization. A gradual spontaneous functional improvement commenced and led several years later to a stabilized state involving moderately severe symptoms. METHODS: In the 15th year of the clinical course, magnetic resonance imaging (MRI) and positron emission tomography (PET) with [(18)F]fluorodeoxyglucose (FDG) and [(11)C] methionine were conducted. Four lines of experiments (clonogenic assay using fibroblasts isolated from a skin biopsy sample of the patient, comet assay, micronucleus assay, and the testing of chromosome aberrations after in vitro irradiation of peripheral blood samples) were performed in a search for an increased individual radiosensitivity. RESULTS: MRI investigations failed to reveal any pathological change. PET demonstrated an increased FDG accumulation, but a negligible [(11)C] methionine uptake in the irradiated spinal cord segments. The radiobiological investigations did not indicate any sign of an increased individual radiosensitivity. CONCLUSIONS: We suggest that the observed partial functional recovery and stabilization of the symptoms of radiogenic LMND may be explained by the higher than normal density of sodium channels expressed along the demyelinated axons of the restored conduction. The increased energy demands of this type of conduction are proved by a higher metabolic rate (increased FDG uptake) of the irradiated spinal cord segments without a substantial regenerative process (lack of detectable protein synthesis).

• Right prefrontal activation produced by arterial baroreceptor stimulation: a PET study.

  Authors: J Weisz, M Emri, J Fent, Z Lengyel, T Márián, G Horváth, P Bogner, L Trón, G Adám

  Neuroreport. 11/2001; 12(15):3233-8.

  This study was performed to test the hypothesis of greater right hemispheric involvement in the processing of baroreceptor stimuli. Carotid sinus baroreceptors were stimulated by rhythmicallyThis study was performed to test the hypothesis of greater right hemispheric involvement in the processing of baroreceptor stimuli. Carotid sinus baroreceptors were stimulated by rhythmically decreasing air pressure in a neck chamber, and under control conditions the thorax was stimulated in a similar manner. Changes in regional cerebral blood flow (rCBF) were measured by PET. Baroreceptor stimulation resulted in rCBF increase in the right anterior-inferior prefrontal cortex (Brodmann areas (BA) 10/44/47) and bilaterally in BA 6/8. We conclude that in at least some stages of baroreceptor information processing the right hemisphere plays a greater role than the left hemisphere.

• [The effect of a single-dose intravenous vinpocetine on brain metabolism in patients with ischemic stroke]

  Authors: B Gulyás, P Bönöczk, A Vas, L Csiba, D Bereczki, I Boros, S Szakáll, L Balkay, M Emri, I Fekete, L Galuska, L Kerényi, S Lehel, T Márián, T Molnár, J Varga, L Trón

  Orvosi hetilap. 04/2001; 142(9):443-9.

  The effect of a single-dose i.v. infusion of vinpocetine on the cerebral blood flow (CBF) and glucose metabolism of post-stroke patients was studied by measuring the regional and global cerebralThe effect of a single-dose i.v. infusion of vinpocetine on the cerebral blood flow (CBF) and glucose metabolism of post-stroke patients was studied by measuring the regional and global cerebral metabolic rates of glucose (CMRglu) and the corresponding kinetic constants before and after treatment. Transcranial Doppler (TCD) and single photon emission tomography (SPECT) measurements were also performed. The cerebral glucose metabolism was significantly higher in the contralateral hemisphere than in the affected one before therapy. In the affected hemisphere the regional glucose metabolism was inhomogenous: relatively low values were measured in the stroke region, whereas it was increased in the peristroke region. Although a single-dose vinpocetine treatment did not affect significantly the regional or global metabolic rates of glucose, the glucose transport (both intracellular up-take and release) was strongly affected in the whole brain, in the contralateral hemisphere and in the peri-infarct area of the symptomatic hemisphere. A slightly increased (not significant, N. S.) cerebral blood flow could be observed in the contralateral and a decreased flow (N. S.) in the symptomatic hemisphere.

• Hypoglycemia activates compensatory mechanism of glucose metabolism of brain.

  Authors: T Márián, L Balkay, I Fekete, Z Lengyel, G Veress, O Esik, L Trón, Z Krasznai

  Acta biologica Hungarica. 02/2001; 52(1):35-45.

  The effect of plasma glucose concentration on the cerebral uptake of [18F]-fluorodeoxy-D-glucose (FDG) was studied in a broad concentration range in a rabbit brain model using dynamic FDG PETThe effect of plasma glucose concentration on the cerebral uptake of [18F]-fluorodeoxy-D-glucose (FDG) was studied in a broad concentration range in a rabbit brain model using dynamic FDG PET measurements. Hypoglycemic and hyperglycemic conditions were maintained by manipulating plasma glucose applying i.v. glucose or insulin load. FDG utilization (K) and cerebral glucose metabolic rate (CGMR) were evaluated in a plasma glucose concentration range between 0.5 mM and 26 mM from the kinetic constant k1, k2, k3 obtained by the Sokoloff model of FDG accumulation. A decreasing set of standard FDG uptake values found with increasing blood glucose concentration was explained by competition between the plasma glucose and the radiopharmacon FDG. A similar trend was observed for the forward kinetic constants k1, and k3 in the entire concentration range studied. The same decreasing tendency of k2 was of a smaller magnitude and was reverted at the lowest glucose concentrations where a pronounced decrease of this backward transport rate constant was detected. Our kinetic data indicate a modulation of the kinetics of carbohydrate metabolism by the blood glucose concentration and report on a special mechanism compensating for the low glucose supply under conditions of extremely low blood glucose level.

• A slow outward current and a hypoosmolality induced anion conductance in embryonic chicken osteoclasts.

  Authors: Z Krasznai, F Weidema, D L Ypey, S Damjanovich, R Gáspár, T Márián

  Acta biologica Hungarica. 01/2001; 52(1):47-61.

  In this paper we report on a hypoosmolality induced current, I(osmo), in embryonic chicken osteoclasts, which could only be studied when blocking a simultaneously active, unidentified slow outwardIn this paper we report on a hypoosmolality induced current, I(osmo), in embryonic chicken osteoclasts, which could only be studied when blocking a simultaneously active, unidentified slow outward current, I(slo). I(slo) was observed in all of the examined cells when both the intracellular and extracellular solutions contained sodium as the major cation and no potassium. The current was outwardly rectifying and activated at membrane potentials more positive than -44 +/- 12 mV (n = 31). The time to half activation of the current was also voltage dependent and was 350 ms at Vm = +80 mV, and 78 ms at Vm = +120 mV. The current did not inactivate during periods up to 5 s. Extracellular 4-AP (5 mM), TEA (5 mM) and Ba2+ (1 mM), blockers of K+ conductances in chicken osteoclasts, did not influence I(slo). However, I(slo) was inhibited by 50 microM extracellular verapamil, which allowed us to study I(osmo) in isolation. Exposure of the osteoclasts to hypotonic solution resulted in the development of a depolarization activated I(osmo). It developed after a 1-min delay and reached its maximum within 10 minutes. Half-maximal activation occurred after 4.4 +/- 0.9 min (n = 9). The current activated within a few ms upon depolarization and did not inactivate during at least 5 sec. I(osmo) reversed around the calculated Nernst potential for Cl- (E(Cl) = +7.3 mV and V(rev) = +5.4 +/- 3.6 mV, n = 9). The underlying conductance, G(osmo) exhibited moderate outward rectification around 0 mV in symmetrical Cl- solutions. Ion substitution experiments showed that G(osmo) is an anion conductance with P(Cl) approximately = P(F) > P(gluc) >> P(Na). I(osmo) was blocked by 0.5 mM SITS but 50 microM verapamil, 5 mM TEA, 5 mM 4-AP, 1 mM Ba2+, 50 microM cytochalasin D and 0.5 mM alendronate did not have any effect on the current. Cl- currents have been implicated in charge neutralization during osteoclastic acid secretion for bone resorption. The present results imply that osmolality may be a factor controlling this charge neutralization.

• Membrane permeability changes induce hyperpolarization in transformed lymphoid cells under high-density culture conditions.

  Authors: T Márián, L Balkay, Z Krasznai, L Trón

  Cytometry. 12/2000; 41(3):186-92.

  BACKGROUND: Membrane potential changes in cells from the human lymphoid B cell line, JY, evoked by increasing cell density in culture were investigated, as data published on other cell types areBACKGROUND: Membrane potential changes in cells from the human lymphoid B cell line, JY, evoked by increasing cell density in culture were investigated, as data published on other cell types are controversial. An attempt was also made to clear the underlying mechanism. METHODS: Nonadherent JY cells were isolated from high-density plateau-phase cultures (type A cells), medium-density log-phase cultures (type B cells), and low-density lag-phase cultures (type C cells). They were analyzed for transmembrane potential, intracellular free concentration of potassium and sodium, membrane permeability for monovalent cations, cell cycle distribution by measuring DNA content, and glucose uptake. RESULTS: C type cells proved to be relatively depolarized (-41 +/- 3 mV) and cells obtained from the highest density cultures hyperpolarized (-60 +/- 3 mV). Intracellular concentrations ([K](i) = 92-97 mM and [Na](i) = 34-35 mM) were almost identical for each type of cell. The sodium/potassium permeability constant ratio in the A and C type of cells was 0.047 and 0.094, respectively. High-density culture conditions resulted in a pronounced G(1)-phase arrest. CONCLUSIONS: Differences in the membrane potential values induced by high-density culture conditions were maintained by changes in the membrane permeability for the monovalent cations.

• Preparation and primary evaluation of [11C]CSC as a possible tracer for mapping adenosine A2A receptors by PET.

  Authors: T Márián, I Boros, Z Lengyel, L Balkay, G Horváth, M Emri, E Sarkadi, A J Szentmiklósi, I Fekete, L Trón

  Applied radiation and isotopes : including data, instrumentation and methods for use in agriculture, industry and medicine. 06/1999; 50(5):887-93.

  A 11C labeled selective adenosine A2A antagonist, (E)-8-(3-chlorostyryl)-1,3-dimethyl-7-[11C]methylxanthine [11C]CSC) was prepared by the reaction of (E)-8-(3-chlorostyryl)-1,3-dimethylxanthine andA 11C labeled selective adenosine A2A antagonist, (E)-8-(3-chlorostyryl)-1,3-dimethyl-7-[11C]methylxanthine [11C]CSC) was prepared by the reaction of (E)-8-(3-chlorostyryl)-1,3-dimethylxanthine and [11C]methyl iodide. The decay-corrected radiochemical yield was 32.3% with a radiochemical purity of 99%, a specific activity of 1.85-5.55 GBq/mumol and a preparation time of 1 h. A primary evaluation of [11C]CSC as a potential tracer for mapping adenosine A2A receptors by positron emission tomography (PET) is also presented. Biodistribution and autoradiographic studies were carried out on Swiss mice and domestic rabbits. In mice the lung showed the highest uptake at 10 min after i.v. injection, followed by the liver, kidney, heart and brain. Inside the brain a high level of radioactivity accumulated in the striatum, in accordance with previous findings on the specific spatial distribution of A2A adenosine receptors and also in the medulla oblongata. Dynamic PET studies on rabbits showed a fast brain uptake of CSC, reaching a maximum in less then 2 min. On the basis of competition experiments with the unlabeled ligand [11C]CSC proves to bind specifically to the appropriate receptor.

• Cerebral effects of a single dose of intravenous vinpocetine in chronic stroke patients: a PET study.

  Authors: S Szakáll, I Boros, L Balkay, M Emri, I Fekete, L Kerényi, S Lehel, T Márián, T Molnár, J Varga, L Galuska, L Trón, D Bereczki, L Csiba, B Gulyás

  Journal of neuroimaging : official journal of the American Society of Neuroimaging. 11/1998; 8(4):197-204.

  The effects of vinpocetine (Cavinton) on the cerebral glucose metabolism of chronic stroke patients are studied with positron emission tomography. The regional and global cerebral metabolic rates ofThe effects of vinpocetine (Cavinton) on the cerebral glucose metabolism of chronic stroke patients are studied with positron emission tomography. The regional and global cerebral metabolic rates of glucose (CMRglu) and the kinetic constants related to them are quantified before and after single-dose intravenous vinpocetine treatment. These measurements are completed with transcranial Doppler sonography and single photon emission computed tomography to explore the possible mechanisms underlying the resulting changes in glucose uptake and metabolism in the brain. The authors' findings indicate that a single-dose vinpocetine treatment, although it does not affect significantly the regional or global metabolic rates of glucose, improves significantly the transport of glucose (both uptake and release) through the blood-brain barrier in the whole brain, the entire contralateral hemisphere, and in the brain tissue around the infarct area of the symptomatic hemisphere. These changes are in accord with increased blood flow in the entire contralateral hemisphere as well as decreased blood flow velocity and increased peripheral vessel resistance in the entire symptomatic hemisphere.

• [Image fusion of different tomographic methods (PET/CT/MRI) effectively contribute to therapy planning]

  Authors: M Emri, O Esik, I Repa, T Márián, L Trón

  Orvosi hetilap. 12/1997; 138(46):2919-24.

  Image fusion (and image registration) is a fundamental topic of three-dimensional (3D) medical image processing. The need to register different imaging modalities arises for several reasons: someImage fusion (and image registration) is a fundamental topic of three-dimensional (3D) medical image processing. The need to register different imaging modalities arises for several reasons: some anatomical details, especially soft tissues, are more easily seen in MRI than CT images, but the bony structures are better visualised by CT. The visualisation of any morphological volume with a functional data set which may come from SPECT or PET is a very important method both for research purposes and for routine diagnostics. Our developed programs allowing us to solve image fusion tasks also contained some free available softwares. These packages helped us to start additional software development for more advanced applications having elaborated this way the informatical background of multimodality image processing. This paper presents three oncological applications of the image fusion: PET-CT-MRI registration for 3D radiotherapy planning, PET-MRI registration in planning surgery and metastasis localization. All these registration processes were solved by the landmark-matching registration method.

• [11C-methionine: an effective radio-tracer for PET scan in the detection of tumors of low proliferating capacity]

  Authors: L Trón, O Esik, Z Kovács, E Sarkadi, L Galuska, L Balkay, M Emri, T Molnár, S Szakáll, E Tóth, T Márián

  Orvosi hetilap. 09/1997; 138(34):2107-12.

  11C-methyl-methionine is available at the PET Center of University Medical School Debrecen since June, 1996. The first 5 oncological examinations were indicated for clinically suspected11C-methyl-methionine is available at the PET Center of University Medical School Debrecen since June, 1996. The first 5 oncological examinations were indicated for clinically suspected recurrent/residual tumorous tissue of low-grade/low proliferative capacity, following the negative or inconclusive results of previous 18F-deoxyglucose (FDG) examinations. In these situations, the methionine examinations provided conclusive results in 4 cases (out of the total of 5 examinations). On the basis of published data and own experience, the authors recommend methionine PET investigations for diagnosis, differential diagnosis and therapy monitoring of tumours of low-grade/low proliferative capacity following inconclusive results of previous FDG examination.

• Flow cytometric determination of intracellular free potassium concentration.

  Authors: L Balkay, T Márián, M Emri, Z Krasznai, L Trón

  Cytometry. 06/1997; 28(1):42-9.

  A flow cytometric assay has been developed for determination of intracellular free potassium concentration ([K+]i). Investigated cells, loaded with the fluorescent pH indicatorA flow cytometric assay has been developed for determination of intracellular free potassium concentration ([K+]i). Investigated cells, loaded with the fluorescent pH indicator 2',7bis-(2-carboxyethyl)-5(and-6)-carboxyfluorescein (BCECF), are incubated in the presence of nigericin, and the intracellular pH is measured. The ionophore maintains the same ratio value [H+]/[K+] on both sides of the cytoplasmic membrane, so [K+]i can be evaluated from the measured intracellular pH (pHi) and the known parameters of the buffer. Application of the method revealed that the intracellular potassium concentration is significantly higher in lymphocytes than in proliferating cells of HUT-78, U266, and JY cell lines. A surprisingly low (60 mM) [K+]i concentration was observed with sperm cells of common carp. This method allows measurements on individual cells, provides data of excellent statistics, and still does not require large amounts of material. These features offer remarkable advantages over other techniques used for intracellular K+ measurements, such as steady-state fluorescence, atom absorption photometry, or energy-dispersive x-ray analysis.

• Role of extracellular and intracellular pH in carp sperm motility and modifications by hyperosmosis of regulation of the Na+/H+ exchanger.

  Authors: T Márián, Z Krasznai, L Balkay, M Emri, L Trón

  Cytometry. 05/1997; 27(4):374-82.

  Recently developed flow cytometric methods (Trón et al.: Mol Immunol 27:1307-1311, 1990) to measure the intracellular pH (pHi) and intracellular potassium concentration in mammalian cells by usingRecently developed flow cytometric methods (Trón et al.: Mol Immunol 27:1307-1311, 1990) to measure the intracellular pH (pHi) and intracellular potassium concentration in mammalian cells by using the fluorescent pH-indicator dye 2',7bis-(2-carboxyethyl)-5 (and-6)-carboxyfluorescein (BCECF) were adopted for measuring these parameters in carp sperm. The intracellular potassium concentration of the carp sperm was 62.4 +/- 5.3 mM. This is very similar to the potassium concentration of the seminal plasma (87 +/- 16 mM), and it suggests a depolarized state of the sperm cell in the semen. An average pHi value of 7.06 +/- 0.11 was obtained by measuring sperm samples taken from ten animals. Changes in the ionic composition of the environment did not alter pHi. Sperm motility was initiated by transferring the cells to an environment of 110 mOsm osmolality. This hypoosmotic shock induced fast changes in the membrane structure that could be reversed by restoring physiologic osmolality. Activation was accompanied by a fast alkalinization of the sperm cells. This pH change was amiloride sensitive, suggesting the involvement of the Na+/H+ exchanger in the activation process. Alkalinization of acid-loaded sperm cells depended on the osmolality of the environment. Equilibrium pHi of these cells in hyperosmotic buffers was substantially lower relative to cells in an isoosmotic environment. Effects of the extracellular and intracellular pH on carp sperm motility were also examined. Extracellular pH below 5.5 abolished sperm motility completely. Alkaline extracellular pH did not alter the duration of sperm motility even at extreme values (pHe = 9.6). Duration of the flagellar motion did not depend on the pHi between values of 6.5 and 8.5; however, it was significantly reduced both below and above this range. No motility was observed below pHi = 6.0 or above pHi = 9.5 with a 10 min incubation time at these pH values prior to activation. Effects of the extracellular and intracellular pH on sperm motility were partially reversible.

• Intracellular pH does not affect drug extrusion by P-glycoprotein.

  Authors: K Goda, L Balkay, T Márián, L Trón, A Aszalós, G Szabó

  Journal of photochemistry and photobiology. B, Biology. 08/1996; 34(2-3):177-82.

  The intracellular pH (pH(i)) of cells exhibiting multidrug resistance (MDR) related to the expression of the P-glycoprotein (Pgp) is often more alkaline than that of the parental cells, as alsoThe intracellular pH (pH(i)) of cells exhibiting multidrug resistance (MDR) related to the expression of the P-glycoprotein (Pgp) is often more alkaline than that of the parental cells, as also observed for the KB-V1/KB-3-1 system in this paper. The possible role of an elevated pH(i) in Pgp-related MDR has been investigated by shifting back the pH(i) of the MDR+ cells to a more acidic value using the mobile proton ionophore carbonylcyanide m-chlorophenylhydrazone (CCCP). The influence of CCCP-evoked delta pH(i) on relative daunorubicin (DNR) accumulation was similar in the case of several Pgp positive and negative cell lines, in view of flow cytometric and radioactive drug accumulation studies and measuring DNR levels in the medium in a flow-through system. Our data argue against a significant effect of pH(i) on Pgp pumping efficiency. However, an indirect connection between pH(i) regulation and the MDR phenotype is suggested by the fact that acidification of the external medium in the presence of verpamil could be observed exclusively in MDR+ cells.

• Flow cytometric determination of absolute membrane potential of cells.

  Authors: Z Krasznai, T Márián, L Balkay, M Emri, L Trón

  Journal of photochemistry and photobiology. B, Biology. 05/1995; 28(1):93-9.

  Membrane potential measurements using fluorescent membrane potential indicator dyes report on relative changes but usually do not result in an absolute value of the measured parameter. The methodMembrane potential measurements using fluorescent membrane potential indicator dyes report on relative changes but usually do not result in an absolute value of the measured parameter. The method developed in this paper is based on the assumption that the negatively charged bis-oxonol distributes across the cytoplasmic membrane according to the Nernst equation. It is further supposed that the fluorescence intensity measured from a given stained cell is a single-value function of the intracellular dye concentration. The protocol suggested incorporates the construction of a calibration curve (fluorescence intensity measured from stained cells vs. extracellular dye concentration). This allows the evaluation of the membrane potential in millivolts using fluorescence readings of the cells both in the depolarized state and in the state of interest. Good agreement was found between absolute membrane potential data of human peripheral blood lymphocytes by our method and results of parallel patch clamp measurements.

• Hypo-osmotic shock induces an osmolality-dependent permeabilization and structural changes in the membrane of carp sperm.

  Authors: T Márián, Z Krasznai, L Balkay, M Balázs, M Emri, L Bene, L Trón

  The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society. 03/1993; 41(2):291-7.

  We carried out spectrofluorimetric and flow cytometric measurements to investigate the effect of hypo-osmotic shock on cell membranes of common carp sperm. The time course of the permeability of theWe carried out spectrofluorimetric and flow cytometric measurements to investigate the effect of hypo-osmotic shock on cell membranes of common carp sperm. The time course of the permeability of the sperm cell membrane, as monitored by DNA-related propidium iodide (PI) fluorescence, was followed for 30 min after dilution of semen in hypo-osmotic environments of different ionic strengths. Spectrofluorimetric measurements indicated a continuous increase in the total PI emission intensity of a sperm suspension. Cell-by-cell flow cytometric measurements suggested that the permeability changes were of the all-or-none type. The permeabilized fraction of cells in the individual samples was time and osmolality dependent. The number and percentage of cells in which DNA was stained by PI increased gradually over time and reached a steady-state plateau value after 5-15 min. This equilibrium fraction of cells with a PI-permeable cytoplasmic membrane displayed an inverse relationship with the osmolality of the diluent, having a near 100% value for fresh water and distilled water. Dilution of sperm in hypo-osmotic medium brought about a fast decrease in the forward light-scattering signal on a short time scale compared to the pre-steady-state time of the permeabilization. With the addition of extracellular Ca2+ (1.8 mM), restoration of the light scattering signal was observed. Permeabilization of the membrane and restoration of light scattering were not coincident in time. We propose a two-dimensional reorganization of the lipid structure as the underlying mechanism of the latter process.

• A novel method for measuring intracellular pH: effect of neutron irradiation on pHi of transformed cells.

  Authors: L Balkay, T Márián, M Emri, L Trón

  Journal of photochemistry and photobiology. B, Biology. 01/1993; 16(3-4):367-75.

  A new flow cytometric method was developed for measuring the intracellular pH (pHi) of mammalian cells using a fluorescent pH indicator dye 2',7-bis-(2-carboxyethyl)-5(and-6)-carboxyfluoresceinA new flow cytometric method was developed for measuring the intracellular pH (pHi) of mammalian cells using a fluorescent pH indicator dye 2',7-bis-(2-carboxyethyl)-5(and-6)-carboxyfluorescein (BCECF). Emission intensities (or their ratios) measured from BCECF-loaded cells can be converted into absolute pHi values using appropriate calibration curves. By comparison of several possible measuring and data evaluation procedures a double-ratio method was suggested as the most advantageous protocol to yield reliable intracellular pH data. This method allows pHi to be determined on a cell-by-cell basis corrected for cell volume and change in geometry of input-output optics of the flow cytometer. Our method applies a standard calibration curve and does not necessitate its reconstruction for each new set of measurements. Cells of the OKT-4 and OKT-8 hybridoma lines were exposed to neutron irradiation of different doses. Irradiated cells underwent a biphasic alkalinization; an instantaneous effect detected within 1.5 h was found to be intensified over 24 h. For the interpretation of data we suggest that the increase in cytoplasmic pH following neutron treatment is evoked by two mechanisms.

• Bretylium-induced voltage-gated sodium current in human lymphocytes.

  Authors: R Gáspár, Z Krasznai, T Márián, L Trón, R Recchioni, M Falasca, F Moroni, C Pieri, S Damjanovich

  Biochimica et biophysica acta. 11/1992; 1137(2):143-7.

  Using the whole-cell variation of the patch-clamp technique it has been determined that 0.25-3 mM bretylium tosylate (BT) exerts a repolarizing effect on partially depolarized human lymphocytes. TheUsing the whole-cell variation of the patch-clamp technique it has been determined that 0.25-3 mM bretylium tosylate (BT) exerts a repolarizing effect on partially depolarized human lymphocytes. The repolarizing effect was ouabain (40 microM)-sensitive, and was inhibited by the removal of external Na+ or by the Na(+)-channel-blocker amiloride (10-44 microM), but K(+)-channel-blockers 4-aminopyridine (0.1-5 mM) and quinine (100 microM) had no effect. The drug induced a sodium dependent, amiloride-sensitive transient inward current reaching its maximum value approx. 20-30 s after the administration of BT and lasting for 6-10 min. This current was activated by depolarization within 25 ms at around -42 mV, its inactivation took about 2 s and its reversal potential was +24 +/- 5 mV. An increase in the intracellular sodium concentration (1.8-3.2 mM) has been observed upon the addition of BT by monitoring the SBFI fluorescence of the dye-loaded cells. It has been shown that whole-cell K+ currents are significantly decreased by BT. The existence of voltage and ligand (BT)-gated sodium channels has been postulated in human lymphocytes. These channels are thought to participate in the initiation of membrane repolarization in human lymphocytes, and thereby influence mitogenic or antigen-induced cell-activation processes.

• Bretylium causes a K(+)-Na+ pump activation that is independent of Na+/H+ exchange in depolarized rat, mouse and human lymphocytes.

  Authors: L Trón, C Pieri, T Márián, L Balkay, M Emri, S Damjanovich

  Molecular immunology. 01/1991; 27(12):1307-11.

  We have studied a bretylium tosylate induced increase of the membrane potentials of partially depolarized rat, mouse and human lymphocytes, using the potential sensitive dye, bis [1,3,We have studied a bretylium tosylate induced increase of the membrane potentials of partially depolarized rat, mouse and human lymphocytes, using the potential sensitive dye, bis [1,3, dibutylbarbituric acid-(5) trimethine oxonol]. The extent of this repolarization is dose-dependent and decreased in magnitude as the temp was reduced from 37 degrees C to room temp. The repolarizing effect is inhibited by K(+)-Na(+)-pump blockers or lack of extracellular Na+. Sodium ion channel blockers are effective in abolishing repolarization only if applied prior to, or simultaneously with, bretylium. Activation of Na+/H+ exchange is not involved in the mechanism of the phenomenon as the latter is completely eliminated in the presence of 10 microM amiloride (concn of the diuretics having no measurable inhibition on the action of the exchanger). These data suggest that bretylium opens ligand- and voltage-gated Na+ channels, and repolarization occurs due to higher activity of the K(+)-Na(+)-pump stimulated by the enhanced intracellular Na+ accumulation.

• Ligand and voltage gated sodium channels may regulate electrogenic pump activity in human, mouse and rat lymphocytes.

  Authors: C Pieri, R Recchioni, F Moroni, L Balkay, T Márián, L Trón, S Damjanovich

  Biochemical and biophysical research communications. 06/1989; 160(3):999-1002.

  Bretylium tosylate - a sodium channel opener - resulted in an increase of membrane potential of depolarized human, rat and mouse T and B lymphocytes. Flow cytometric membrane potential measurementsBretylium tosylate - a sodium channel opener - resulted in an increase of membrane potential of depolarized human, rat and mouse T and B lymphocytes. Flow cytometric membrane potential measurements with bis-oxonol revealed that the above hyperpolarizing effect was amiloride, ouabain, tetrodotoxin, azide and temperature sensitive. The effect showed an absolute dependence on the extracellular sodium but it was insensitive to the extracellular Ca2+ level. The voltage gating of the effect can be eliminated by either an increase of the extracellular potassium concentration or low doses of veratrin. The existence of a voltage and ligand gated sodium channel is suggested in the plasma membrane of all kinds of lymphocytes. The hyperpolarization is explained by an increased activity of the electrogenic sodium-potassium ATP-ase. Induced opening of such sodium channels may regulate the electrogenic pump activity and indirectly cell activation.