Alister G. Macdonald

The adaptation of biological membranes to temperature and pressure: Fish from the deep and cold

The homoeostatic regulation of bilayer order is a property of functional importance. Arguably, it is best studied in those organisms which experience and must overcome disturbances in bilayer order which may be imposed by variations in temperature of hydrostatic pressure. This article reviews our recent work on the adaptations of order in brain membranes of those fish which acclimate to seasonal changes in temperature or which have evolved in extreme thermal or abyssal habitats. The effects of temperature and pressure upon hydrocarbon order and phase state are reviewed to indicate the magnitude of the disturbances experienced by animals in their environments over the seasonal or evolutionary timescale. Acclimation of fish to altered temperature leads to a partial correction of order, while comparison of fish from extreme cold environments with those from temperate or tropical waters reveals a more complete adaptation. Fish from the deep sea also display adaptations of bilayer order which largely overcome the ordering effects of pressure.

A dilatometric investigation of the effects of general anaesthetics, alcohols and hydrostatic pressure on the phase transition in smectic mesophases of dipalmitoyl phosphatidylcholine

Abstract The effect of general anaesthetics, alcohols and hydrostatic pressure on the thermal transition in dipalmitoyl phosphatidylcholine multilayer liposomes has been measured using dilatometry. The volume increasse at the transition ( ΔV t ) is 0.0350 ± 0.0003 ml/g. the transition temperature ( T t ) 41.84 ± 0.09°C and the width of the transition 1.025 ± 0.18°C. ΔH calculated by the Clapeyron-Clausius equation is 8.4 kcal/mol. The n -alcohols C 3 C 5 reduced the transition temperature without affecting the transition width which was however, increased by n -hexanol. Trichloroethylene, the fluorescent probe N -phenyl-1-naphthyl-amine, and methoxyflurane all increased the transition width (reduced the cooperativity of the transition) with a simultaneous depression of T t . Methoxyflurane caused a two-stage transition expansion. Diethyl ethers effect has similarities with both the C 3 and C 6 alcohols. Generally ΔV t was unaffected by the agents. Pressure increased T t by 0.0238°C/atm linearly over the range 1–300 atm in both treated and untreated liposomes, and therefore cannot be said to antagonize anaesthetics. In both treated and untreated liposomes ΔV t and the width of the transition were unaffected by pressure. Pressure thus reverses the effects of anaesthetics on T t but not their spread of the transition width.

Homeoviscous adaptation under pressure. III. The fatty acid composition of liver mitochondrial phospholipids of deep-sea fish

Abstract Homeoviscous adaptation of biological membranes to high hydrostatic pressure has been investigated by determining the differences in lipid composition of membranes from fish obtained from depths between 200 and 4000 m. The fatty acid composition of phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine/inositol and cardiolipin from a liver mitochondrial fraction was analysed by capillary gas-liquid chromatography. The ratio of saturated to unsaturated fatty acids significantly and negatively related to depth in PC and PE as predicted by homeoviscous adaptation to pressure. Thus, deep sea species possess greater proportions of unsaturated fatty acids than shallow species. Cardiolipin showed the opposite trend. An unsaturation index was not significantly related to depth in any phospholipid fraction.

The effects of nitrous oxide on a glutamate-gated ion channel and their reversal by high pressure; a single channel analysis

Nitrous oxide reversibly affects the kinetics, but not the conductance, of the qGluR channel of locust muscle. 0.5 atm N2O at 20.5 degrees C was without effect but both 1.5 and 2.7 atm significantly reduced the probability of the channel opening, the frequency of opening and the mean open time, and prolonged the mean closed time. 100 atm helium was without effect on these parameters, but when 98.5 atm He was combined with 1.5 atm N2O they, and the associated dwell time distributions, were restored to normal. 100 atm similarly combined with 2.7 atm N2O exerted a comparable trend which fell short of significance. The results are consistent with nitrous oxide binding to the channel with a significant molar volume increase, which pressure opposes. This suggests that nitrous oxide may cause conformational changes in the channel, and that the pressure reversal of nitrous oxide anaesthesia in animals could be caused by molecular antagonism.

Temperature, pressure and cholesterol effects on bilayer fluidity. A comparison of pyrene excimer/monomer ratios with the steady-state fluorescence polarization of diphenylhexatriene in liposomes and microsomes

Pyrene excimer/monomer (E/M) ratios have been compared with the steady-state fluorescence polarization (P) of diphenylhexatriene (DPH) in multilamellar liposomes of dilaurylphosphatidylcholine and rat liver microsomes. The purpose was to use the well-understood properties of DPH to reveal the nature of bilayer fluidity which pyrene manifests as an E/M ratio. Reducing the temperature (from 37 degrees C to 8 degrees C), increasing the hydrostatic pressure (from 0.1 to 70 MPa), and, in liposomes, cholesterol enrichment (up to 0.30 mole fraction) separately decreased the E/M ratios and increased P. The pyrene membrane/buffer partition coefficient was affected by temperature but not by pressure, and in the case of cholesterol enrichment, it was assumed to be unaffected. Plots of P as a function of the E/M ratio showed the two to be closely correlated (r = 0.99 in liposomes and 0.96 in microsomes), independent of the treatment used to reduce fluidity. The apparent activation volume and enthalpy for excimer formation was calculated and compared with published data. Pyrene E/M ratios probably reflect the intermolecular volume (fluidity) of the outer region of the bilayer, which is reduced by a decrease in temperature and an increase in pressure and cholesterol. DPH reports the bilayer interior, which is similarly ordered by the experimental treatments. The regional distinction between the two probes, however, accounts for the divergence of E/M ratios and P, which has been reported in membranes enriched with fluidizing fatty acids.

Homeoviscous adaptation under pressure: the pressure dependence of membrane order in brain myelin membranes of deep-sea fish

Steady-state and time-resolved anistropy of 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescence have been used to compare the hydrocarbon order of brain myelin membranes from a shallow water (plaice) and two deep-sea fish species (Coryphenoides rupestris and Coryphenoides armatus). At atmospheric pressure the deep sea fish displayed lower steady-state anisotropies than shallow water species although the pressure dependence of anisotropy was similar in all species. Time-resolved measurements allowed the separate determination of the rate of probe motion from the amplitude of that motion. Anisotropy decays were analysed in terms of two correlation times and a constant (r infinity). The r infinity and mean value of P2 order parameter for all species increased with pressure, the graphs for deep-sea species being translated to higher pressures relative to shallow-water species. The resulting pressure coefficients for C. armatus was distinctly less than for the two shallower species. These time-resolved studies show that the interspecific differences provide for similar order parameters in all three species when corrected to their respective habitat conditions of pressure and temperature. This indicates that myelin order is highly conserved despite the profound ordering effects of high hydrostatic pressure.

Effects of high pressure on the channel gated by the quisqualate-sensitive glutamate receptor of locust muscle and its blockade by ketamine; a single-channel analysis

The effects of high pressure on the channel gating kinetics of the quisqualate-sensitive L-glutamate receptor (qGluR) of locust muscle have been investigated using a megaohm seal patch-clamp technique. Pressure was applied with helium gas and recordings were carried out at 20.5 degrees C with Rb+ as the main charge-carrying cation in the patch pipette. The mean open time of the qGluR channel was unaffected by 10 and 30 MPa, but it was significantly reduced at 50 MPa. A high proportion of brief openings (mean 0.808 ms) was seen at 50 MPa but not at lesser pressures. Also, in contrast to lesser pressures, 50 MPa prolonged the mean closed time and reduced both the frequency and probability of channel opening. 10(-6) M ketamine significantly reduced the mean channel open time, as previously reported. A pressure of 10 MPa which alone had no effect on the qGluR channel, restored the mean open time in the presence of 10(-6) M ketamine to the value obtained in the absence of the anaesthetic. This implies the shortening of qGluR channel open time by ketamine involves a large + delta V and, therefore, probably conformational changes in the channel. However 10 MPa did not restore the distribution of open times to normal.

Experiments on ion channels at high pressure

Ion channels are distinctive membrane proteins which provide a gated pathway for diffusing ions. High pressure (<100 MPa) affects the kinetics of gating but not the conductance of the channel. Dynamic structural studies of channels at high pressure are, thus far, conspicuously absent but functional properties are studied at the single channel level with the patch clamp technique.

Effects of High Pressure on Cellular Processes

Publisher Summary This chapter discusses that high hydrostatic pressure affects equilibria and reaction rates through the molar volume changes involved in the reaction system as a whole. The chapter explains how high hydrostatic pressure affects molecules of biological interest, and their interactions; the ways in which pressure affects cellular processes; and how animals, including humans are affected by pressure. Along with high pressure, the chapter also intends to stimulate interest in a particular approach to physiology. The discussion on molecular effects of pressure and temperature includes equilibrium processes in aqueous solution, lipid bilayers under pressure and rates of chemical reactions. It is mentioned that hydrostatic pressure is not directional and that osmotic pressure is also quite separate from hydrostatic pressure. Hydrostatic pressure is a force acting in all directions, in the air one breathes, in water, or in body fluids. While high pressure, surprisingly perhaps, dissociates molecules in aqueous solution, it has the intuitively expected effect on lipid bilayers, compressing and ordering their structure. The thermodynamics of equilibrium processes in aqueous solution, and the phase state of lipid bilayers at high pressure, inevitably fall short of explaining how pressure affects rates of reaction. Muscle, a tissue whose cells are conspicuously filled with polymerized proteins, is affected by high pressure in ways determined by the contraction cycle. Simple processes, such as the diffusion of gases or water in aqueous solution, are little affected by the pressure range of interest here. Chondrocytes are cells, which synthesize the load-bearing extracellular matrixthat covers the articulating surfaces of joints. The buoyancy of aquatic animals subjected to significant pressure is interesting because buoyancy requires the creation of a void or region of low-density, working against the ambient pressure. Animals and bacteria live at high pressures in the deep sea, down to the greatest depths where the pressure is approximately 100 MPa. Several technological applications have arisen from high pressure biology—the manipulation of ploidy, and of membrane proteins; sterilization and inactivation of microorganisms; food processing; and safe human diving.

Aspects of Eukaryotic Cells

The first experiments in high pressure physiology were observations on aquatic animals and isolated tissues, such as muscle and nerve (Regnard 1885). The interpretation of the effects seen had to await the development of modern physiology and biochemistry, and are still far from complete. In its second stage of development the field became more biophysical, with research into (a) bioluminescence (Johnson et al. 1974); (b) the colloidal structure of eukaryotic cells (Marsland 1970); and (c) the excitability of nerves and the contractility of muscle (Cattell 1936). These investigations progressed relatively rapidly because in (a), rate process theory and enzyme kinetics, and in (b) colloidal ideas on the dynamic structure of cytoplasm, each provided a rationale for interpreting the results. Excitability and contractility remained complex phenomenological fields, strong in experimental work but lacking the molecular details necessary for applying physical chemical methods of analysis. This situation has now changed as is clear from sections 3 and 4. One of the pioneers in the field, Ebbecke, clearly perceived the possibilities of the subject noting that “we can expect too, the compression effects which universally affect physiological processes to be instructive in the dynamics and molecular kinetics of normal life and cell exchange” (Ebbecke 1936). Furthermore, Ebbecke appeared to have ideas about the fluid mosaic nature of cell membranes, well ahead of his time (see Macdonald 1987, for a translated account). In the relatively recent growth of our understanding of the properties of lipid bilayers and protein-lipid interactions, high pressure has played its role but that is not dealt with here, however, having been reviewed recently (Macdonald 1987; Wong et al. 1988).