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Biomedical subjects

I C Smith

Publications and source records attributed to I C Smith.

At least 127 records · Page 7Linked to original sources
PubMed1986

Changes in miniature end-plate potentials due to moderate hypertonicity at the frog neuromuscular junction.

The effect of an increase in tonicity on the amplitude and frequency of miniature end-plate potentials (m.e.p.p.s) following superfusion of 50 mM- or 100 mM-sucrose-Ringer solution was determined from intracellular recording at end-plates of the frog cutaneous pectoris muscle, in the absence of drugs and using on-line statistical analysis. An immediate decrease in mean amplitude of the order of 20% control was associated with a marked increase in mean frequency. A delayed increase in mean amplitude, independent of frequency, followed the initial response in the majority of end-plates exposed to 100 mM-sucrose-Ringer solution. The net gain was of the order of 34% and was attained over 20-40 min. This was not reversible over at least 20 min. There was an increase in the variability of m.e.p.p. amplitudes associated with the increase of mean amplitude but this was reversible. A moderate increase in tonicity is considered to induce two kinds of presynaptic changes. First, an immediate increase in the probability of release of smaller quantal packets and secondly, an increased loading of transmitter into vesicles within the readily releasable store.

Action Potentials↗
PubMed1986

Mevalonic acid is partially synthesized from amino acids in Halobacterium cutirubrum: a 13C nuclear magnetic resonance study.

13C nuclear magnetic resonance revealed an unusual pathway for the biosynthesis of lipids in Halobacterium cutirubrum and H. halobium. Mevalonic acid was not synthesized from three acetyl-coenzyme A molecules, as has been suggested previously, and the branch-methyl and methine carbons in phytanyl chains were derived from neither acetate nor glycerol. Instead, they were supplied by the degradation of amino acids, in particular of lysine. Presumably, two different types of two-carbon fragments were used simultaneously by halobacteria for the biosynthesis of mevalonate. The labeling pattern of squalene supported the above conclusions. Based on these data, a general scheme is proposed to account for the contribution of lysine-to-lipid biosynthesis.

Acetates↗
PubMed1986

Large-scale selection synchrony of Tetrahymena thermophila.

A method is described, based on the phagocytosis of colloidal ferrite particles, which gives highly synchronous populations of Tetrahymena thermophila. To ensure a successful synchrony, the cell culture doubling time, the limits of the phagocytic period and the distribution of cell stages must first be determined. Once these parameters are known, synchrony can be achieved under a variety of growth conditions and with cultures ranging in volume from a few millilitres to 12 litres or more. The main advantages of the method are that the apparatus required is simple, large volumes of cells can be handled easily, and the synchronous populations can be prepared within a few hours. In principle, the method should be applicable to any cell population in which phagocytosis occurs discontinuously over the cell cycle.

Animals↗
PubMed1986

Microcomputers and biologists.

Microcomputers have now become a working tool of the biologist. This article explores some of the consequences of this revolution both in terms of the skills and working habits needed and of the range of uses which have been found. The primary consequence is that biologists must learn how to become computer managers. This does not mean learning BASIC and computer architecture, though this will usually prove valuable, but that we must be able to grasp from the flood of sales literature and computer specialists what tasks can or should be undertaken with the aid of microcomputers. Secondly it means being able to choose and maintain the right machinery for the tasks needed. An overview of applications shows four main areas of use. 1) The standard packages where one can expect the computer manufacturer to provide a complete working system. These are word-processing, database reference systems, spreadsheet calculations, graphics production and communications. 2) Specialist scientific programs such as statistics, data processing and model making. 3) The use of the microcomputer as a laboratory instrument for control or measurement. 4) Educational applications. Each of these areas has separate, often conflicting, requirements which can only be balanced by the user.

Audiovisual Aids↗
PubMed1985

Biological antioxidants.

The mechanism of lipid peroxidation and the ways in which the rate of this reaction can be reduced by small quantities of certain specific chemicals, called antioxidants, are described. The types and roles of the different antioxidants found in living systems are considered. Vitamin E (alpha-tocopherol) has long been recognized as an important lipid-soluble, chain-breaking antioxidant. It has an unexpectedly high reactivity towards peroxyl radicals, which can be understood only after detailed consideration of its structure. It is the major antioxidant of its class in human blood and its effectiveness in plasma is greatly improved by a synergistic interaction with water-soluble reducing agents such as ascorbic acid. Experiments designed to locate vitamin E within phospholipid bilayers and to discover the origin of the different biopotencies of stereoisomers of alpha-tocopherol are also described.

Antioxidants↗
PubMed1985

Estimation of the location of natural alpha-tocopherol in lipid bilayers by 13C-NMR spectroscopy.

Natural, 2R,4R',8R'-alpha-tocopherol (vitamin E), labelled selectively with 13C in the methyl group at position 5, was incorporated into unilamellar vesicles of egg phosphatidylcholine. The vesicles are impermeable to the shift reagent Pr3+ and, in the presence of this reagent, separate 13C resonances due to labelled alpha-tocopherol in the outer and inner monolayers could be observed with relative intensities, 2:1. Subsequent addition of the relaxation reagent Gd2+ causes broadening and greatly shortened spin-lattice relaxation times for the resonance due to alpha-tocopherol in the outer monolayer only. These data confirm that alpha-tocopherol is located in both halves of the bilayers with its more hydrophilic chroman moiety very near the lipid-water interface, and indicate that the methyl group at position 5 of the alpha-tocopherol in the inner monolayer must be at least 40 A from the aqueous interface of the outer monolayer.

Lipid Bilayers↗
PubMed1985

Acyl chain dynamics of phosphatidylethanolamines containing oleic acid and dihydrosterculic acid: 2H NMR relaxation studies.

The dynamical behavior of the acyl chains of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine, and 1-palmitoyl-2-dihydrosterculoyl-sn-glycero-3-phosphoethanolamine has been investigated by using 2H T1 and T2 relaxation times. Lipids were labeled at the 5-,9-,10-, and 16-positions of the sn-2 acyl chain. The profile of deuterium spin-lattice relaxation rate (T1(-1) vs. chain position is characterized in all systems by a marked discontinuity at the positions of the carbon-carbon double bond and the cyclopropane ring; the deuterons at these positions have relaxation rates which are greater than at any other labeled position of the sn-2 chain. For both types of sn-2 acyl chain, assuming a single-exponential correlation time and that the motion is within the rapid regime, the phosphatidylcholine lipid systems are less mobile than their phosphatidylethanolamine analogues. Systems containing an oleoyl chain are more dynamic than their analogues containing a dihydrosterculoyl chain. The rates of motion of the sn-2 acyl chains of phosphatidylethanolamine in a bilayer structure are slower than those of the lipid in an inverted hexagonal structure. In the hexagonal phase, the motional rates of a dihydrosterculoyl chain are slower than those of the corresponding positions of an oleoyl chain.

Fatty Acids↗
PubMed1985

31P nuclear magnetic resonance studies of crayfish (Orconectes virilis). The use of inversion spin transfer to monitor enzyme kinetics in vivo.

31P nuclear-magnetic resonance (NMR) has been used to observed in vivo the steady-state levels of phosphorus-containing metabolites in the crayfish Orconectes virilis and the intracellular pH of the abdominal muscle was determined. Measurement of spin-lattice relaxation times and spin transfer experiments have enabled calculation of unidirectional rate constants and activation energies for the arginine kinase reaction in vivo.

Adenosine Triphosphate↗
PubMed1985

2H NMR evidence for antibiotic-induced cholesterol immobilization in biological model membranes.

The interaction of the polyene antibiotic filipin with membrane sterols has been studied by deuterium nuclear magnetic resonance of the molecular probes [2,2,3,4,4,6-2H6]cholesterol and 1-myristoyl-2-[4',4',14',14',14'-2H5]myristoyl-sn-glycero-3-phospho- choline. At physiological temperatures, there is evidence of filipin-induced cholesterol immobilization in the membrane. The 2H NMR spectra of cholesterol show two domains in which ordering and dynamics are very different. In one of these, cholesterol is static on the 2H NMR time scale, whereas in the other it undergoes rapid axially symmetric motions similar to those it exhibits in the drug-free membrane; this indicates that the jumping frequency of cholesterol between the labile and immobilized domains is less than 10(5) s-1. The distribution of cholesterol between these two sites is temperature dependent; at 0 degrees C all sterol molecules are immobilized, whereas at 60 degrees C they are almost totally in the labile site. In contrast to cholesterol, the phospholipids sense only one type of environment, at both the top and center of the bilayer, indicating that cholesterol acts as a screen, preventing the lipids from direct interaction with the antibiotic. At low temperature, the ordering of the lipid in the presence of cholesterol does not change upon filipin addition, whereas at elevated temperatures the local ordering of both the lipid and the labile cholesterol is significantly lower than that in the absence of the drug. Moreover, there is a very important difference between the degree of local ordering as measured by the lipids and by cholesterol at high temperatures.(ABSTRACT TRUNCATED AT 250 WORDS)

Cholesterol↗
PubMed1985

Effects of replacement of a double bond by a cyclopropane ring in phosphatidylethanolamines: a 2H NMR study of phase transitions and molecular organization.

The thermotropic behavior and molecular properties of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE) and 1-palmitoyl-2-dihydrosterculoyl-sn-glycero-3-phosphoethanolamine (PDSPE) have been investigated by 2H NMR spectroscopy using samples selectively labeled at the 5'-, 9'-, 10'-, and 16'-positions of the sn-2 chains. Comparison with the corresponding phosphocholine analogues (POPC and PDSPC), obtained as intermediate synthetic products, was used to monitor the role of the polar head group. Replacement of the choline moiety by ethanolamine increased the gel to liquid-crystal transition temperature by 10-32 degrees C and led to a significantly higher ordering of the fatty acyl chains in the liquid-crystalline bilayer state. The lateral compression effect, due to the smaller area per polar head group in PE, results in a bilayer to hexagonal phase transition at elevated temperatures. The effects on both PC and PE due to replacement of the olefinic group by a cyclopropane unit are similar. A decrease in the temperature of the gel to liquid-crystal phase transition, Tc, is observed upon introduction of a cyclopropane ring; it goes from 26 degrees C in POPE to approximately 10 degrees C in PDSPE. In addition, a very significant broadening of the transition profile is observed. These observations are consistent with the poor packing ability of mixed saturated and cyclopropane-containing chains due to the bulky substituent effect. The temperature of the bilayer-hexagonal phase transition of PE samples was decreased by 15-20 degrees C on replacement of oleoyl chains by dihydrosterculoyl chains at the sn-2 position.(ABSTRACT TRUNCATED AT 250 WORDS)

Calorimetry, Differential Scanning↗
PubMed1985

A versatile perfusion technique for metabolic studies by NMR.

A perfusion technique applicable to metabolic studies by NMR is described in detail. The utility and versatility of the approach are demonstrated by following gluconeogenesis from [2-13C]pyruvate in the perfused, isolated mouse liver, and lipogenesis from [2-13C]acetate in perfused Acanthamoeba castellanii cells embedded in agarose filaments.

Acetates↗
PubMed1985

Plasmodium berghei: gluconeogenesis in the infected mouse liver studied by 13C nuclear magnetic resonance.

Using 13C nuclear magnetic resonance, we have compared the gluconeogenic activity of perfused livers isolated from normal starved mice and mice highly parasitized with Plasmodium berghei, using [2-13C]pyruvate as substrate. In both types of livers, 13C labeling of glucose carbons occurred in positions 1, 2, 5, and 6. The equal proportions of [1,6-13C]- and [2,5-13C]glucose in livers from malarial and normal mice suggests that pyruvate enters the gluconeogenic pathway directly and, to an equal extent, via the tricarboxylic acid cycle. The normalized signal heights indicated that at a given time after the addition of [2-13C]pyruvate the degree of 13C labeling in glucose carbons was reduced in livers from malarial animals, when compared to livers from normal animals. During the course of the perfusion experiment, the [2-13C]lactate resonance signal was always more intense from livers of malarial animals than from normal animals. A reduced activity of hepatic gluconeogenesis in malarial animals was further confirmed by a separate set of perfusion experiments which showed a 56% reduction of the measured rate of glucose production in livers from malarial animals, with respect to that of normal animals. A lowered NAD/NADH ratio in livers from malarial animals would explain the increased proportion of lactate observed in the spectra and be related to a decreased gluconeogenic rate. A more reduced oxidoreduction level in the hepatocytes of a malarial animal would result from a defect in the oxidative phosphorylation activity of mitochondria.

Animals↗
PubMed1984

Interaction of amphotericin B with membrane lipids as viewed by 2H-NMR.

The effects of amphotericin B upon the organization and dynamics of multibilayer membranes of dimyristoylphosphatidylcholine (DMPC) were investigated by means of 2H-NMR. At high amphotericin B concentrations (30 mol% with respect to the lipid) and at temperatures above 25 degrees C, DMPC experiences two different environments which are in slow exchange on the 2H-NMR time scale. In one of these, the lipid is immobilized by the antibiotic, in a molar ratio of approximately 1:1, whereas the lipid unsequestered by amphotericin B is more ordered than in its pure state. This ordering effect is perceived at relatively low antibiotic doses (4%). The local lipid order, and the relative percentage, of sequestered DMPC, are temperature-independent (up to 65 degrees C), whereas the ordering of the unsequestered lipid domain is not. The perturbation induced by amphotericin B is manifest similarly at the edges as well as in the center of the bilayer. Antibiotic addition leads to large decreases in the transverse relaxation time, T2, of the labelled lipid, but not in the spin-lattice relaxation time, T1. This indicates an increased density of slow motional modes and little change in rapid motions.

Amphotericin B↗
PubMed1984

Amphotericin and model membranes. The effect of amphotericin B on cholesterol-containing systems as viewed by 2H-NMR.

The interactions between amphotericin B and sterol-containing model membranes were monitored by 2H-NMR of deuterium-labelled dimyristoylphosphatidylcholine (DMPC), cholesterol and epicholesterol. The addition of amphotericin B to a cholesterol/DMPC (3:7) system was perceived differently by the lipid, depending upon the depth in the bilayer: no structural change was manifest in the acyl chain region associated with the plateau in molecular ordering (C4'), whereas the lipid clearly senses two environments near the center of the bilayer (C13', C14'). The amount as well as the ordering properties of the more ordered antibiotic-induced component, sensed at C14', increased with decreasing temperature. The structural parameters of deuterium-labelled cholesterol in cholesterol/DMPC mixtures were unchanged upon addition of amphotericin B, regardless of the bilayer depth. Upon addition of amphotericin B, the lipid T1 values are unchanged, whereas the T2 values are reduced by a factor of 2. The minimum in T1 observed for cholesterol in DMPC at 32-35 degrees C was shifted towards 38-40 degrees C in the presence of amphotericin B. Epicholesterol-containing dispersions of DMPC had properties similar to those of their cholesterol-containing analogs; a noticeable difference between the systems was an approx. 10% increase in the segmental order parameters on the addition of amphotericin B to the system containing the alpha-isomer of cholesterol. The concept of a dynamic complexation between amphotericin B and sterol is discussed.

Amphotericin B↗
PubMed1984

31P-NMR studies of the freeze-tolerant larvae of the gall fly, Eurosta solidaginis.

31P NMR was applied to an examination of the freeze-tolerant larvae of the gall fly, Eurosta solidaginis. Resonances from sugar phosphates, inorganic phosphate, adenylates and arginine phosphate were identified. Two peaks of Pi were identified corresponding to intracellular and extracellular Pi. Anoxia produced an expected decrease in peak intensities of ATP and arginine phosphate while the peak of intracellular Pi was enhanced and shifted to indicate intracellular acidification during anoxia. Spectra of whole larvae were monitored over a temperature range from -30 degrees to +25 degrees C. No abrupt alterations in the spectra were seen at the point of extracellular freezing which occurs at about -8 degrees C but temperature had dramatic effects upon the peak intensities of ATP and arginine phosphate. A reversible increase/decrease in peak intensities, relative to Pi, was observed as temperature was raised/lowered. At 15 degrees and -20 degrees C, the beta peak of ATP was 64% and 2% of the peak intensity of Pi while that of arginine phosphate was 78% and 11%, respectively. This temperature effect was not an artifact of instrumentation (as model solutions containing Pi, ATP and arginine phosphate did not show this effect) or a result of changes in the total amounts of these compounds in the cell with temperature. Rather it is apparent that these molecules become restricted in their rotational movement as temperature is lowered perhaps via binding to subcellular components. Changes in the amounts of freely soluble ATP and arginine phosphate with temperature could have important implications for metabolism and its control. Analysis of the effect of temperature on the chemical shift of Pi was also used to determine pH in the intracellular and extracellular compartments. Temperature change had no effect on extracellular (hemolymph) pH which remained constant at 6.1-6.3. Intracellular pH varied with temperature, however, from pH 6.8 at 15 degrees C to pH 7.3 at -12 degrees C with a change, delta pH/delta 0, of -0.0185 degrees C consistent with alphastat regulation.

Adenosine Triphosphate↗
PubMed1984

Plasmodium berghei: electron spin resonance and lipid analysis of infected mouse erythrocyte membranes.

Red blood cells from mice infected with Plasmodium berghei and from uninfected mice were labeled with stable, free radical derivatives of stearic acid. Electron spin resonance spectra of these samples showed that the degree of molecular order in these membranes decreased, and the rate of motion of the probe increased, with increasing levels of parasitemia. Parasitemia increased the ratio of unsaturated to saturated 18-carbon fatty acids, and decreased the percentage of arachidonic acid and of cholesterol. The effects of parasitemia on the membrane properties correlated with decreases in cholesterol/fatty acid ratios.

Animals↗
PubMed1984

NMR methods for characterizing the state of the surfaces of complex mammalian cells.

It is shown that narrow 1H NMR resonances may be observed in cancer cells, and that these belong to fatty acyl chains of membrane lipids. A variety of NMR techniques such as Gaussian-Lorentzian deconvolution, and T1 and T2 measurements, may be used to subdivide these resonances further. The results of these various methods require that in the membrane structures the observed lipids tumble isotropically and sufficiently rapidly to give motionally narrowed 1H NMR lines.

Animals↗