Biomedical subjects

F Riemann

Publications and source records attributed to F Riemann.

11 recordsLinked to original sources

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↗

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↗

PubMed1975

Preparation of erythrocyte ghosts by dielectric breakdown of the cell membrane.

Dielectric breakdown of membranes of red blood cells was observed in high electric fields (approx. 10-3-10-4 V/cm) using an improved Coulter Counter with hydrodynamic focussing. In making measurements of the size distributions of red blood cells as a function of increasing electric field strength it was found that a sharp discontinuity occurred in the otherwise linear relation between the pulse heights in the Coulter Counter and the electric field strength due to dielectric breakdown of the membranes. Solution of Laplace's equation for the electric field generated at breakdown in the cell membranes yeilds a mean value of about 1.6 V. for the membrane potential of red blood cells. Due to the dielectric break-down, release of hemoglobin occurred. Mechanical rupture of the red blood cells by the hydrodynamic forces in the orifice of the Coulter Counter or thermal rupture could be excluded as hemolysing mechanisms. The leaky ghost cells resealed at 37 degrees C. as shown by incorporation of 131I-labeled albumin and repeated dielctric breakdown.

Animals↗

PubMed1974

Dielectric breakdown of cell membranes.

With human and bovine red blood cells and Escherichia coli B, dielectric breakdown of cell membranes could be demonstrated using a Coulter Counter (AEG-Telefunken, Ulm, West Germany) with a hydrodynamic focusing orifice. In making measurements of the size distributions of red blood cells and bacteria versus increasing electric field strength and plotting the pulse heights versus the electric field strength, a sharp bend in the otherwise linear curve is observed due to the dielectric breakdown of the membranes. Solution of Laplace's equation for the electric field generated yields a value of about 1.6 V for the membrane potential at which dielectric breakdown occurs with modal volumes of red blood cells and bacteria. The same value is also calculated for red blood cells by applying the capacitor spring model of Crowley (1973. Biophys. J. 13:711). The corresponding electric field strength generated in the membrane at breakdown is of the order of 4 . 10(6) V/cm and, therefore, comparable with the breakdown voltages for bilayers of most oils. The critical detector voltage for breakdown depends on the volume of the cells. The volume-dependence predicted by Laplace theory with the assumption that the potential generated across the membrane is independent of volume, could be verified experimentally. Due to dielectric breakdown the red blood cells lose hemoglobin completely. This phenomenon was used to study dielectric breakdown of red blood cells in a homogeneous electric field between two flat platinum electrodes. The electric field was applied by discharging a high voltage storage capacitor via a spark gap. The calculated value of the membrane potential generated to produce dielectric breakdown in the homogeneous field is of the same order as found by means of the Coulter Counter. This indicates that mechanical rupture of the red blood cells by the hydrodynamic forces in the orifice of the Coulter Counter could also be excluded as a hemolysing mechanism. The detector voltage (or the electric field strength in the orifice) depends on the membrane composition (or the intrinsic membrane potential) as revealed by measuring the critical voltage in E. coli B harvested from the logarithmic and stationary growth phases. The critical detector voltage increased by about 30% for a given volume on reaching the stationary growth phase.

Animals↗

PubMed1974