The State of Water in Human and Dog Red Cell Membranes

The apparent activation energy for the water diffusion permeability coefficient, P_d, across the red cell membrane has been found to be 4.9 ± 0.3 kcal/mole in the dog and 6.0 ± 0.2 kcal/mole in the human being over the temperature range, 7° to 37°C. The apparent activation energy for the hydraulic conductivity, L_p, in dog red cells has been found to be 3.7 ± 0.4 kcal/mole and in human red cells, 3.3 ± 0.4 kcal/mole over the same temperature range. The product of L_p and the bulk viscosity of water, η, was independent of temperature for both dog and man which indicates that the geometry of the red cell membrane is not temperature-sensitive over our experimental temperature range in either species. In the case of the dog, the apparent activation energy for diffusion is the same as that for self-diffusion of water, 4.6–4.8 kcal/mole, which indicates that the process of water diffusion across the dog red cell membrane is the same as that in free solution. The slightly, but significantly, higher activation energy for water diffusion in human red cells is consonant with water-membrane interaction in the narrower equivalent pores characteristic of these cells. The observation that the apparent activation energy for hydraulic conductivity is less than that for water diffusion across the red cell membrane is characteristic of viscous flow and suggests that the flow of water across the membranes of these red cells under an osmotic pressure gradient is a viscous process.     

Effect of Geometrical and Chemical Constraints on Water Flux across Artificial Membranes

Studies have been made on the temperature dependence of both the hydraulic conductivity, L_p, and the THO diffusion coefficient, ω, for a series of cellulose acetate membranes (CA) of varying porosity. A similar study was also made of a much less polar cellulose triacetate membrane (CTA). The apparent activation energies, E_a, for diffusion across CA membranes vary with porosity, being 7.8 kcal/mole for the nonporous membrane and 5.5 kcal/mole for the most porous one. E_a for diffusion across the less polar CTA membrane is smaller than E_a for the CA membrane of equivalent porosity. Classical viscous flow, in which the hydraulic conductivity is inversely related to bulk water viscosity, has been demonstrated across membranes with very small equivalent pores. Water-membrane interactions, which depend upon both chemical and geometrical factors are of particular importance in diffusion. The implication of these findings for the interpretation of water permeability experiments across biological membranes is discussed.     

Thermal inactivation and stabilization of lysozyme substrate-- Micrococcus lysodeicticus cells

Heat inactivation of the acetonic powder of Micrococcus lysodeicticus cells suspended in phosphate buffer pH 6.2 was quantitatively characterized in the temperature range from 34 to 52 degrees. The total value of the rate constant for heat inactivation of the cells equals 2.88 X 10(8) exp(-18360/RT) sec-1. The activation parameters of the process at 34 degrees are the following: delta H* = 17.7 kcal/mole; delta S* = 21.8 E. U.; delta F* = 24.4 kcal/mole. The effect of ethylene glycol, mannitol, dextran, polyvinyl alcohol (PVA) and polyethylene glycols with different molecular weights on the lysis rate and cell stability was studied. Polyvinyl alcohol was found to be the most effective stabilizer. At concentrations of about 10(-5) it enhances the thermostability of the cells threefold.     

Heats of activation for the exosmotic flow of water across the membrane of leucocytes and leukemic cells

The permeability of the membranes of different types of leucocytes from several mammalian species to the exosmotic flow of water has been measured at different temperatures. Heats of activation for the process have been calculated. Results indicate that the leucocyte has an unusually high heat of activation for the osmotic movement of water across its outer membrane, ranging between 13 to 18 kcal/mole. Unconstrained water movement usually has values between 3 to 5 kcal/mole. This property characterizes all leucocytes examined to date, regardless of species or type. It is apparent that water is intimately associated with the outer boundary of the cell and that this association is quite sensitive to the structural changes which must occur in response to changes in temperature.     

Temperature-dependent kinetics of the activities of influenza virus

The temperature dependence of membrane interactions between PR8 influenza virus and virus receptor (GD1a)-containing liposomes was studied. For quantitation, the octadecylrhodamine B chloride (R18) membrane marker was incorporated into liposomes at quenched concentrations. Upon interaction with target membranes, the marker gets diluted, and dequenching can be measured in a fluorescence spectrophotometer. Rate constants were calculated from the dequenching curves under low pH conditions, which allow for fusion, and at neutral pH, where no specific fusion occurs. Activation energies were determined from Arrhenius plots. The results were compared with the temperature dependence of other viral activities like infectivity, hemolysis, and fusion with erythrocytes. For the slow reaction at pH 7.4, where only non-specific lipid transfer takes place, the activation energy was about 24 kcal/mole between 15 degrees C and 45 degrees C. For the fast, hemagglutinin (HA)-specific fusion reaction (pH 5.3), a very low activation energy (approximately 7 kcal/mole) was found between 25 degrees C and 37 degrees C, whereas below 25 degrees C it was much higher (approximately 34 kcal/mole). The temperature range with low activation energy coincides with the one for optimal infectivity, hemolysis, and fusion with erythrocytes. Furthermore, it is the same range in which the conformational change of HA takes place, which in the absence of a partner membrane leads to an irreversible inactivation of the fusion protein.     

Mathematical modelling of corneal swelling

This paper presents a differential model of the corneal transport system capable of modelling thickness changes in response to osmotic perturbations applied to either limiting membrane. The work is directed towards understanding corneal behaviour in vivo. The model considers the coupled viscous flows within the corneal stroma and across the epithelial and endothelial membranes. The flows within the stroma are established based on transport theory in porous media, while the flows across the membranes are described using the phenomenological equations of irreversible thermodynamics. The ability of the numerical model to reproduce corneal thickness changes in response to endothelial perturbations was tested against available experimental data. The sensitivity of the model to changes in stromal and membrane transport coefficients was examined.     

Temperature Dependence of Vasopressin Action on the Toad Bladder

Toad bladders were challenged with vasopressin at one temperature, fixed on the mucosa with 1% glutaraldehyde, and then subjected to an osmotic gradient at another temperature. Thus, the temperature dependence of vasopressin action on membrane permeability was distinguished from the temperature dependence of osmotic water flux. As the temperature was raised from 20° to 38°C, there was a substantial increase in the velocity of vasopressin action, but osmotic flux was hardly affected. In this range of temperature the apparent energy of activation for net water movement across the bladder amounted to only 1.2 kcal/mole, a value well below the activation energy for bulk water viscosity. It is suggested that osmotic water flux takes place through narrow, nonpolar channels in the membrane. When the temperature was raised from 4° to 20°C, both vasopressin action as well as osmotic water flux were markedly enhanced. Activation energies for net water movement were now 8.5 kcal/mole (4°–9°C) and 4.1 kcal/mole (9°–20°C), indicating that the components of the aqueous channel undergo conformational changes as the temperature is lowered from 20°C. At 43°C bladder reactivity to vasopressin was lost, and irreversible changes in selective permeability were observed. The apparent energy of activation for net water movement across the denatured membrane was 6.6 kcal/mole. Approximately 1 µosmol of NaCl was exchanged for 1 µl of H₂O across the denatured membrane.     

Temperature dependence of ultrasound-induced cell killing: The role of membrane fluidity

Chinese hamster cells in suspension were exposed to 20 kHz ultrasound (US) at 54 W/cm² and various temperatures between 2 and 44 °C. Activation energies were 2.6 and 24 kcal/mole below and above 35 °C, respectively. Procaine, a local anaesthetic drug known to increase membrane fluidity, enhanced cellular inactivation by US above 41 °C, increasing the activation energy to 62 kcal/mole. The inactivation of the bacterium Salmonella typhimurium by US was also dependent on the exposure temperature, with an activation energy of 2.9 kcal/mole between 2 and 44 °C. These data are most simply explained by the hypothesis that membranes are a major target for cellular inactivation by US and that the fluidity of the membranes is important in this respect.     

The influence of gramicidin A and valinomycin of the permeability of mammalian erythrocytes.

1. The K+ transport in erythrocytes induced by gramicidin A or valinomycin is a first-order reaction. The activation energy of the induced transport is low and amounts to 6 and 10 kcal/mole, respectively. The indirect method for calculation of the driving force of diffusion, c, is given; in pig erythrocytes the c value for gramicidin A is 21.7, and for valinomycin 35.4 mM-KCl. 2. The valinomycin-induced transport was found to be the same in erythrocytes and biomolecular lipid membranes. The gramicidin A-induced transport corresponds to that of a single gramicidin channel, and not to the multichannel transport observed in the model systems. 3. Erythrocytes of various mammals show large differences in sensitivity to the ionophores. No correlation could be found between membrane lipids and the induced permeability. The role of membrane proteins in ionophore-induced permeability is discussed.     

Effects of temperature and pH on the water exchange through erythrocyte membranes: Nuclear magnetic resonance studies

Summary The temperature and pH dependence of water exchange has been studied on isolated erythrocytes suspended in isotonic buffered solutions. At pH 7.4 a break in the Arrhenius plot of water exchange time at around 26°C was found. The mean value of the apparent activation energy of the water exchange time at temperatures higher than that of the discontinuity was 5.7 kcal/mole (±0.4); at lower temperatures the values of the apparent activation energy were below 1.4 kcal/mole. The pH dependence of water exchange time of isolated erythrocytes revealed a marked increase of the water exchange time values in the acid range of pH; a much smaller variation of the same parameter occurs between pH 7.0 and 8.0. These finding could be correlated with other processes involving erythrocyte membranes that showed similar pH and temperature dependence and were considered to indicate state transitions in the membranes. It is suggested that the temperature and pH effects on water diffusion indicate that conformational changes and cooperative effects are implicated in the mechanism of this transport process.Institute for Isotopic and Molecular Technology.     

Multiple membrane systems as biological models

Summary The current-voltage equations for double, triple, and quadruple membrane systems are derived in closed form from the flow equations of irreversible thermodynamics. Numerical examples show that the behavior of these systems is very similar to that of nerve and muscle membranes. Multiple membrane systems exhibit resting potentials which do not have a characteristic Nernst concentration dependence; nonpermeant ions play a significant role in this nonlogarithmic behavior. Furthermore, multiple membrane systems have rectification properties similar to those of biological membranes. The direction of rectification is determined by the polarity of the membrane systems, not by the ionic concentrations in the bathing solutions.     

Membrane Transport Generated by the Osmotic and Hydrostatic Pressure. Correlation Relation for Parameters L p, σ, and ω

Standard approach to membrane transport generated by osmotic andhydrostatic pressures, developed by Kedem and Katchalsky, is based onprinciples of thermodynamics of irreversible processes. In this paper wepropose an alternative technique. We derive transport equations from fewfairly natural assumptions and a mechanistic interpretation of the flows.In particular we postulate that a sieve-type membrane permeability isdetermined by the pore sizes and these are random within certain range.Assuming that an individual pore is either permeable or impermeable tosolute molecules, the membrane reflection coefficient depends on the ratioof permeable and impermeable pores. Considering flows through permeableand impermeable pores separately, we derive equations for the total volumeflux, solute flux and the solvent flux across the membrane. Comparing themechanistic equations to the Kedem-Katchalsky equations we find the formereasier to interpret physically. Based on the mechanistic equations we alsoderive a correlation relation for the membrane transport parameters L _p,, and . This relation eliminates the need for experimentaldetermination of all three phenomenological parameters, which in somecases met with considerable difficulties.     

Fast flip-flop of cholesterol and fatty acids in membranes: implications for membrane transport proteins

PURPOSE OF REVIEW: The rates by which unesterified fatty acids and cholesterol move through and desorb from membranes have been difficult to measure, in part because of the simple structures of these lipids but also because methods have generally not clearly distinguished the two steps of membrane transport. Lack of definitive knowledge has given rise to speculation about the mechanism(s) of membrane 'transport' proteins for fatty acids and cholesterol. RECENT FINDINGS: New biophysical and biochemical approaches have provided evidence that fatty acids and cholesterol exhibit rapid diffusion (flip-flop), as fast as milliseconds, across both protein-free phospholipid bilayers and cell membranes. In contrast, desorption of the cholesterol molecule from a membrane surface (hours) is much slower than that of common dietary fatty acids (milliseconds to seconds). SUMMARY: Knowledge of these properties provides a framework for understanding transport and metabolism of cholesterol and fatty acids and how their putative membrane and intracellular transporters might function.     

Subversion of membrane transport pathways by vacuolar pathogens

Mammalian phagocytes control bacterial infections effectively through phagocytosis, the process by which particles engulfed at the cell surface are transported to lysosomes for destruction. However, intracellular pathogens have evolved mechanisms to avoid this fate. Many bacterial pathogens use specialized secretion systems to deliver proteins into host cells that subvert signaling pathways controlling membrane transport. These bacterial effectors modulate the function of proteins that regulate membrane transport and alter the phospholipid content of membranes. Elucidating the biochemical function of these effectors has provided a greater understanding of how bacteria control membrane transport to create a replicative niche within the host and provided insight into the regulation of membrane transport in eukaryotic cells.     

The permeability of liposomes to nonelectrolytes. I. Activation energies for permeation.

The effect of temperature on the permeability of nonelectrolytes across liposomal membranes above and below their transition temperature has been studied by using an osmotic method. Below their transition temperature, liposomes are osmotically insensitive structures but, on addition of gramicidin A, the water permeability so increased that the permeability of solutes could be studied. The measured activation energies for permeation of a variety of nonelectrolytes has been found to increase when a) there is an increase in the capability of the solutes to form hydrogen bonds in water, b) the cholesterol concentration in the membranes increases and c) the membranes pass from a liquid-crystalline to a solid-crystalline state. The change in the activation energy for permeation per hydrogen bond is about 1.8 kcal/mole for all the different liposome systems investigated; the only solute tested that deviated from this correlation was urea, whose activation energy for permeation across a gramicidin-containing system was much lower than expected from its hydrogen-bonding capacity. This finding suggests that urea is permeating across the gramicidin pore. Although the literature contains only incomplete data relating the activation energies for permeation of nonelectrolytes across biological membranes to their hydrogen-bonding capacity, the available evidence suggests that there is a similar correlation to that found in liposomes. Thus, the average increase in the activation energy per hydrogen bond for permeation across ox red cell membranes (Jacobs, Glassman & Parpart, J. Cell. Comp. Physiol. 7:197, 1935) is 2.2 plus or minus 0.4 kcal/mole, a value that is similar to that obtained in liposomes. However, the activation energies for water and urea are - in such a system - very much lower than expected, suggesting that they, too, are permeating by some parallel route such as an aqueous pore.     

Über die Nachrichtenaufnahme durch biologische Receptoren

Summary The Hodgkin-Huxley theory of ion fluxes across membranes during excitation is extended to explain the graduated depolarisation (receptor potential) of sensory cell membranes. Electric circuit equivalents of living membranes are developed. Driving forces and velocity coefficients are represented by means of electric parameters. From this model active and passive ionic fluxes can be calculated quantitatively on the basis of transport equations derived from irreversible thermodynamics. Thus the circuit equivalent may be used as an analog computer. Electric receptor models allow a reproduction of all potential curves which have been derived in electrophysiological experiments on PD-receptors. The results obtained by the use of this model agree with the results obtained in biological experiments under various conditions of stimuli. The significance of solution compartments for intra-and extracellular ions in relation to the time functions of various conditions are discussed in detail. This models are of heuristic value in experimental research. In combination with neuron networks they can be used for the analysis of information theoretical problems.     

Ion homeostasis, channels, and transporters: an update on cellular mechanisms

The steady-state maintenance of highly asymmetric concentrations of the major inorganic cations and anions is a major function of both plasma membranes and the membranes of intracellular organelles. Homeostatic regulation of these ionic gradients is critical for most functions. Due to their charge, the movements of ions across biological membranes necessarily involves facilitation by intrinsic membrane transport proteins. The functional characterization and categorization of membrane transport proteins was a major focus of cell physiological research from the 1950s through the 1980s. On the basis of these functional analyses, ion transport proteins were broadly divided into two classes: channels and carrier-type transporters (which include exchangers, cotransporters, and ATP-driven ion pumps). Beginning in the mid-1980s, these functional analyses of ion transport and homeostasis were complemented by the cloning of genes encoding many ion channels and transporter proteins. Comparison of the predicted primary amino acid sequences and structures of functionally similar ion transport proteins facilitated their grouping within families and superfamilies of structurally related membrane proteins. Postgenomics research in ion transport biology increasingly involves two powerful approaches. One involves elucidation of the molecular structures, at the atomic level in some cases, of model ion transport proteins. The second uses the tools of cell biology to explore the cell-specific function or subcellular localization of ion transport proteins. This review will describe how these approaches have provided new, and sometimes surprising, insights regarding four major questions in current ion transporter research. 1) What are the fundamental differences between ion channels and ion transporters? 2) How does the interaction of an ion transport protein with so-called adapter proteins affect its subcellular localization or regulation by various intracellular signal transduction pathways? 3) How does the specific lipid composition of the local membrane microenvironment modulate the function of an ion transport protein? 4) How can the basic functional properties of a ubiquitously expressed ion transport protein vary depending on the cell type in which it is expressed?     

Maturation of membrane function: The permeability of the rat erythroblastic leukemic cell to water and to non-electrolytes

The erythroblastic leukemia produced in Long-Evans rats by the administration of 7, 8, 12 trimethylbenz (a) anthracene has been used as a model of the most immature form of the erythrocyte series. In conjunction with studies of the maturation of several other membrane functions, the permeability of this cell to water and to certain definitive non-electrolytes was measured with osmotic methods. The hydraulic conductivity, L_p was 6.2 micro (minute)⁻¹, (atm)⁻¹ at 25°C, quite high and characteristic of mature erythrocytes, but different from values of 0.65 for immature myeloid cells. The effect of temperature provided an energy of activation of 4.4 kCal/mole, also typical of mature mammalian erythrocytes but again different from 13 to 18 kCal/mole for immature myeloid cells. Urea was compared to thiourea. The permeability coefficient for urea was 76.7 micra (minute)⁻¹ ± 13.8 (S. E.); the value for thiourea was 1.55 micra (minute)⁻¹ ± 0.18 (S. E.). Phloretin at 0.25 mM inhibited urea permeability by 90% with 50% inhibition occurring at 0.05 mM. Inhibition was reversible. Permeability to the glycols was also compatible with mature erythrocytes. We infer from these findings that the structure which underlies these basic, passive membrane functions is laid down early and persists after loss of nucleus and subsequent maturation.