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Biomedical subjects

U Zimmermann

Publications and source records attributed to U Zimmermann.

At least 271 records · Page 15Linked to original sources
PubMed1978

Pressure probe technique for measuring water relations of cells in higher plants.

A new method is described for continuously measuring cell turgor pressure (P), hydraulic conductivity (L(p)), and volumetric elastic modulus (epsilon) in higher plant cells, using a pressure probe. This technique permits volume changes, DeltaV, and turgor pressure changes, DeltaP, to be determined with an accuracy of 10(-5) to 10(-6) mul and 3 to 5.10(-2) bar, respectively.The main principle of the new method is the same as the pressure probe developed by Zimmermann and Steudle in which pressure is transmitted to a pressure transducer by means of an oil-filled capillary introduced into the cell. In order to use the pressure probe for small tissue cells, the effective compressible volume of the apparatus has to be sufficiently small in comparison to the volume of the cell itself. This is achieved by accurately fixing the oil/cell sap boundary in the very tip of the microcapillary by means of an electronic feedback mechanism, so that the effective volume of the apparatus is reduced to about 2 to 10% of the cell volume. In this way also, errors arising from compressibility of the apparatus and temperature fluctuations can be excluded.Measurements on tissues cells of Capsicum annuum fruits yield epsilon values of 2 to 25 bar. Furthermore, epsilon can be shown to be a function of both cell turgor pressure and cell volume; epsilon increases with increasing turgor pressure and is higher in larger cells.

Journal Article↗
PubMed1978

Penetration and entrapment of large particles in erythrocytes by electrical breakdown techniques.

Human erythrocytes suspended in isotonic solutions were subjected to haemolysis by application of an electric field pulse to the cell suspension. The field strengths used were 12 and 16kV/cm, respectively; the pulse duration 40 microseconds. The lysed cells showed resealing properties. The permeability change of the membrane generated by the field pulse and by the subsequent osmotic processes were large enough to facilitate the penetration and entrapment of ferritin and Latex particles (diameter: 0.091 and 0.176 micron, respectively) as revealed by electron microscopy. Correct identification of the Latex particles in the electron-micrographs indicated that LOYTER et al. [J. Cell Biol. 66, 292 (1975)], who recently demonstrated the entrapment of Latex spheres in erythrocytes prepared by osmotic haemolysis mistook electron-dense bodies probably consisting of denaturated protein for Latex particles. Under conditions of osmotic haemolysis, carried out according to BODEMANN and PASSOW, particles could only occasionally be detected within the membrane itself and never within the cell interior, suggesting that the electrical haemolysis method is much more effective in the generation of large holes in the membrane.

Cell Membrane Permeability↗
PubMed1977

Preparation of uniform haemoglobin free human erythrocyte ghosts in isotonic solution.

A method is described for the preparation of haemoglobin free human erythrocyte ghosts in isotonic solutions using dielectric breakdown technique. In this single haemolytic procedure, almost complete removal of haemoglobin (less than or equal to 0.1%) was achieved by subjecting the erythrocytes suspended in phosphate buffered, isotonic KCl solution at 0 degrees C to three consecutive electrical field pulses of 16 kV/cm in the presence of 10 mM EDTA; EDTA was used to prevent electrical haemolysis. Haemolysis is induced by subsequent dilution with isotonic and isoionic solution to lower the EDTA concentration. Haemolysis is complete after 5 min; the cells are centrifuged, washed and resuspended in a solution of the same composition and osmolarity containing 4 mM MgCl2, but no EDTA. The resealing process, carried out at 37 degrees C, was complete in about 1 h. Measurements of the size distribution of the ghost cells in the hydrodynamically focusing Coulter Counter at varying field strengths in the orifice revealed that the ghost population is nearly uniform. The mean (modal) volume of the ghost cells was 110--120 micronm3 when suspended in phosphate buffered NaCl solution. The apparent breakdown voltage was about 1.3 V.

Erythrocyte Membrane↗
PubMed1977

The effect of pressure on the electrical breakdown in the membranes of Valonia utricularis.

The interpretation of electrical breakdown in terms of electro-mechanical instabilities, predicts that the breakdown potential should decrease with increasing cell turgor pressure. Experiments were conducted to test this hypothesis on cells of Valonia utricularis over a turgor pressure range of 0.5-10(5)-5.0-10(5) N/m2. Electrical breakdown was measured using intracellular electrodes and 500 mus current pulses. The pressure was monitored by an intracellular micropipette pressure transducer. The results obtained show a linear decrease in the critical breakdown potential with pressure. The effective compressive modulus of the cell membrane, gamma, is calculated from the slope of this line to 69+/-10-10(5) N/m2 (average value of seven measurements). This is consistent with the theoretical prediction of the electromechanical model using our previously determined values of the elastic modulus of the membrane. A theoretical analysis is given of the effects of pressure on the breakdown, This includes also considerations of the indirect effect of pressure on the membrane via stretching of the cell wall with a possible coupling of such strains to the cell membrane. The results and analysis presented allow us to conclude on the basis of the experimentally determined breakdown P.K. of 959 mV that the region of membrane where electrical breakdown occurs is a dielectric with one of the following combinations of parameters: (A) a thickness delta=7-9 nm with a dielectric constant epsilon=greater than 10, e.g. a hydrated protein spanning the whole membrane. (B) delta=4-5 nm with epsilon=3-8, e.g. a lipoprotein of lipid bilayer dimensions. (C) delta approximately 2 nm with epsilon=2-3, e.g. a half lipid bilayer. If we assume that the breakdown P.D. of the tonoplast and plasmalemma are identical, that is 480 mV, then there is only one reasonable choice for the membrane thickness and the dielectric constant: delta=2 nm, epsilon=3-8, e.g. a (lipo) proteinaceous module facing a half life lipid bilayer.

Cell Membrane↗
PubMed1977

Effect of turgor pressure and cell size on the wall elasticity of plant cells.

Direct measurements of the volumetric elastic modulus, in, of cells of a higher plant were performed on the epidermal bladder cells of Mesembryanthemum crystallinum using a pressure probe technique. Measurements on giant algal cells (Valonia, Nitellopsis) are given for comparison. Giant celled algae and M. crystallinum bladders have elastic moduli, in, which depend strongly on turgor pressure, P, and on cell volume, V. The in values of Mesembryanthemum bladders range between 5 bar at zero pressure and 100 bar at full turgor pressure (3-4 bar). in increased with cell size (volume) at a given turgor pressure, and this volume dependence was pronounced more in the high pressure range. From the in (P) characteristics, complete volume-pressure curves were obtained for Mesembryanthemum bladders and giant algal cells. The results suggest that the in (P) and in (V) characteristics of all plant cells are similar. The significance of the pressure and volume effects for the water relations and growth processes of plant cells is discussed briefly.

Journal Article↗
PubMed1976

Electrical hemolysis of human and bovine red blood cells.

The external electric field strength required for electrical hemolysis of human red blood cells depends sensitively on the composition of the external medium. In isotonic NaCl und KCl solutions the onset of electrical hemolysis is observed at 4 kV per cm and 50 per cent hemolysis at 6 kV per cm, whereas increasing concentrations of phosphate, sulphate, sucrose, inulin and EDTA shift the onset and the 50 per cent hemolysis-value to higher field strengths. The most pronounced effect is observed for inulin and EDTA. In the presence of these substances the threshold value of the electric field strength is shifted to 14 kV per cm. This is in contrast to the dielectric breakdown voltage of human red blood cells which is unaltered by these substances and was measured to be approximately 1 V corresponding in the electrolytical discharge chamber to an external electric field strength of 2 to 3 kV per cm. On the other hand, dielectric breakdown of bovine red blood cell membranes occurs in NaCl solution at 4 to 5 kV per cm and is coupled directly with hemoglobin release. The electrical hemolysis of cells of this species is unaffected by the above substances with exception of inulin. Inulin suppressed the electrical hemolysis up to 15 kV per cm. The data can be explained by the assumption that the reflection coefficients of the membranes of these two species to bivalent anions and uncharged molecules are field-dependent to a different extent. This explanation implies that electrical hemolysis is a secondary process of osmotic nature induced by the reversible permeability change of the membrane (dielectric breakdown) in response to an electric field. This view is supported by the observation that the mean volumes of ghost cells obtained by electrical hemolysis can be changed by changing the external phosphate concentration during hemolysis and resealing, or by subjecting the cells to a transient osmotic stress immediately after the electrical hemolysis step. An interesting finding is that the breakdown voltage, although constant throughout each normally distributed ghost size distribution, increases with increasing mean volume of the ghost populations.

Animals↗
PubMed1976

Enzyme loading of electrically homogeneous human red blood cell ghosts prepared by dielelctric breakdown.

Human red blood cell ghosts were prepared by electrical haemolysis at 0 degrees C in isotonic solutions using a discharge chamber which was part of a high voltage circuit. The size distribution of the ghosts was normally distributed, the modal (=mean) volume was approx. 115 mum3, performing the electrical haemolysis in the following solution: 105 mM KCI, 20 mM NaCL, 4mM MgCl2, 7.6 mM Na2HPO4, 2.94 mM NaH2PO4, 10 mM glucose, pH 7.2. Resealing was carried out at o degrees C for 10 min (after the haemolytic step) and then for further 20 min at 37 degrees C. The mean volume of the ghost preparation could be changed by variation of the phosphate concentration in the above solution replacing a part of NaCl by phosphate (5 mM phosphate: 94 mum3, 15 mM phosphate: 135 mum3). The breakdown voltage of the ghost cell membranes measured with a hydrodynamic focusing Coulter Counter depends on the mean volume (94 mum3 = 1.04 V, 134 mum3 = 1.36 V). On the other hand, the breakdown voltage is constant throughout each size distribution pointing to an "electrically homogeneous" ghost preparation. The sensitiviity of the Coulter Counter to detect electrical inhomogeneities in the membranes of a ghost population is demonstrated by dielectric breakdown measurements of an apparently normally distributed ghost preparation containing two different "electrically homogeneous" ghost population i.e. with two different breakdown voltages. The ghost cells obtained by electrical haemolysis in the above solution containing 10mM phosphate were fairly impermeable to sucrose and behave like an ideal osometer. It is further demonstrated that ghost cells can be loaded with enzymes (e.g. urease) and drugs using this technique and that these loaded ghost cells can be used as bioactive capsules for clinical application.

Biological Transport↗
PubMed1976

Turgor Pressure Regulation in Valonia utricularis: Effect of Cell Wall Elasticity and Auxin.

The electrical membrane resistance rho(0) of the marine alga Valonia utricularis shows a marked maximum in dependence on the turgor pressure. The critical pressure, P(c), at which the maximum occurs, as well as its absolute value, rho(0) (max), are strongly volume-dependent. Both P(c) and rho(0) (max), increase with decreasing cell volume. It seems likely, that these relationships reflect the elastic properties of the cell wall, because the volumetric elastic modulus, epsilon, is also volume-dependent, increasing hyperbolically with cell volume. Both P(c) and rho(0) (max) can be affected by external application of indole-3-acetic acid at concentrations of 2.10(-7)m to 2 .10(-5)m. The critical pressure is shifted by 1.2 to 6 bars toward higher pressures and the maximum membrane resistance increased up to 5.6-fold. During the course of the experiments (up to 4 hours), however, IAA had no effect on the volumetric elastic modulus, epsilon.The maximum in membrane resistance is discussed in terms of a pressure-dependent change of potassium fluxes. The volume dependence of P(c) and rho(0) (max) suggests that not only turgor pressure but also epsilon must be considered as a regulating parameter during turgor pressure regulation. On this basis a hypothesis is presented for the transformation of both, a pressure signal and of changes in the elastic properties of the cell wall into alterations of ion fluxes. It is assumed that the combined effects of tension and compression of the membranes as well as the interaction between membrane and cell wall opposingly change the number of transport sites for K(+) providing a turgor-sensing mechanism that regulates ion fluxes. The IAA effects demonstrated are consistent with this view, suggesting that the basic mechanisms for turgor pressure regulation and growth regulation are similar.Any relation connecting growth rate with turgor pressure should be governed by two parameters, i.e. by a yielding pressure, at which cell growth starts, and by the critical pressure, at which it ceases again.

Journal Article↗
PubMed1975

Dielectric breakdown measurements of human and bovine erythrocyte membranes using benzyl alcohol as a probe molecule.

Dielectric breakdown of intact erythrocytes and subsequent haemolysis in the presence of increasing concentrations of benzyl alcohol were investigated by means of an electrolytical discharge chamber and a hydrodynamic focusing Coulter Counter. Low concentrations of the drug stabilized human and bovine erythrocytes against haemolysis induced by dielectric breakdown of the cell membrane in isotonic solutions, while high concentrations caused lysis similar to hypotonic and mechanical haemolysis. The stabilizing effect of the drug on electrically induced haemolysis depends on the pulse length of the applied electric field. The critical dielectric breakdown voltage of the membranes of intact cells decreases progressively with increasing benzyl alcohol concentrations, at which the membrane is also more stabilized against electrical and osmotic haemolysis. Occasionally, an increase in the dielectric breakdown voltage is observed at drug concentrations at which lysis occurs. A similar depedence of the breakdown voltage on drug concentration was found for human erythrocyte ghost cells prepared by dielectric breakdown. The results are consistent with the electromechanical model suggested for the dielectric breakdwon mechanism and with the assumption of Metcalfe, using NMR and ESR techniques, that the fluidity of the membrane increases with increasing benzyl alcohol concentration.

Animals↗
PubMed1975

Release and uptake of haemoglobin and ions in red blood cells induced by dielectric breakdown.

External electric field strengths of the order of 10-3 minus10-4 v-cm-minus1 induce potassium release and concomitant sodium uptake in human and bovine red blood cells, as demonstrated in an electrolytic discharge chamber. The reversible increase of the membrane permeability once the critical membrane potential is reached is caused by dielectric breakdown of the membrane. The values of the critical membrane potential differences calculated from the potassium release and sodium uptake curves are close to those which were calculated from dielectric breakdown measurements in a hydrodynamic focussing Coulter Counter using the Laplace equation. With bovine red blood cells, the potassium release and the concomitant sodium uptake is coupled with haemoglobin release from the cells, while with human red blood cells much higher external electric field strengths are required for haemoglobin release. The external electric field strength required for solute release and uptake in bovine and human red blood cells depends on the pulse length, particularly below a value of about 10 mus, when a strong increase in the field strength occurs with decreasing pulse lengths. At 50-100 mus pulse lengths an asymptotic value of the critical electrical field strength of 2.6 kV-cm-minus1 for the modal volume of human red blood cells and 2.8 kV-cm-minus1 for the modal volume of bovine red blood cells is reached, corresponding to a critical membrane potential difference of about 1.1 V for both species. This value is close to that measured directly for dielectric breakdown of the membranes of Valonia utricularis (0.85 V, 20 degrees C). The increase in electric field strength with decreasing pulse length can be explained by the capacitance of the membrane, which becomes the rate limiting step for the temporal build-up of the electric potential across the membrane. The time constant of this process was determined to be approx. 10 mus. The critical membrane potential difference for breakdown is therefore pulse-length independent. The breakdown of the membrane can be interpreted by an electromechanical collapse of the membrane material. Numerical considerations of the dynamics of this membrane collapse predict that the breakdown time is a very rapid process.

Animals↗