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D F Wilson

Publications and source records attributed to D F Wilson.

At least 217 records · Page 12Linked to original sources
PubMed1982

Adenine nucleotide efflux in mitochondria induced by inorganic pyrophosphate.

The efflux of mitochondrial adenine nucleotide which is induced by addition of PPi to suspensions of rat liver mitochondria has been investigated. This efflux of adenine nucleotide is greatly stimulated by the uncoupler FCCP at 1 microM, Vmax being 6.7 nmol/min per mg protein as compared to 2.0 nmol/min per mg protein in its absence. The depletion process is inhibited by carboxyatractyloside. The Km for PPi of 1.25 mM is essentially unchanged when uncoupler is added. Quantitation of the individual adenine nucleotide species (ATP, ADP and AMP) and their relationship to the rate of efflux suggests that ADP is the predominant species being exchanged for PPi.

Adenine Nucleotides↗
PubMed1982

Mitochondrial transmembrane pH and electrical gradients: evaluation of their energy relationships with respiratory rate and adenosine 5'-triphosphate synthesis.

Mitochondria from rat liver were suspended in ionic media, and parallel measurements were made of the respiratory rate, intramitochondrial volumes, and the phosphorylation state [[ATP]/([ADP][Pi])], as well as transmembrane pH gradients (from the distribution of labeled weak acids and bases) and electrical gradients (from the distribution of labeled lipid-soluble ions and of K+ in the presence of valinomycin). A decrease in the pH gradient was observed with additions of sodium propionate, acetate, or lactate. In the case of propionate, the pH gradient decreased from -0.94 +/- 0.10 (control) to -0.09 +/- 0.08 unit (with 3 mM propionate) without change in any of the other measured parameters. Variation of the intramitochondrial volumes was achieved by changing [K+] (+valinomycin) and by diluting the choline chloride media. The transmembrane electrical potential decreased from -150 mV under control conditions to approximately -75 mV in a 70 mosM medium or when 8 mM K+ (+valinomycin) was present. The changes in [ATP]/([ADP][Pi]) were as follows: from 1.8 X 10(4) (control) to 3.9 X 10(2) M-1 with 8 mM K+ (+valinomycin) or 1.0 X 10(4) M-1 when the osmolarity was decreased from 300 to 70 mosM. This corresponds to a decrease in the energy utilized for ATP synthesis from 14.1 to 11.9 kcal/mol (from 58.9 to 49.7 kJ/mol) with added K+ and to 13.8 kcal/mol (57.7 kJ/mol) with decreased osmolarity. No direct correlation was found between the changes in transmembrane electrical potential, pH gradient, or total proton electrochemical gradient and either the respiratory rate or [ATP]/([ADP][Pi]).

Adenosine Triphosphate↗
PubMed1982

Regulation of intracellular pH by human peripheral blood lymphocytes as measured by 19F NMR.

We have measured the intracellular pH of human peripheral blood lymphocytes by means of high-resolution 19F NMR spectroscopy using D,L-2-amino-3,3-difluoro-2-methylpropanoic acid (F2MeAla) as a probe. Lymphocytes readily took up the methyl ester of F2MeAla, and endogenous esterase hydrolyzed the ester to the free amino acid inside the cell. This alpha-methyl amino acid is not metabolized by the cell, and its 19F NMR spectrum exhibits large pH-dependent shifts as the alpha-amino group is protonated. The size of the 19F shifts, the high sensitivity of 19F NMR, and the favorable pKa of the alpha-amino group of F2MeAla (pKa = 7.3) allowed us to measure intracellular pH of lymphocytes at 25-30 degrees C with approximately 5-min acquisition times. Measurements at various external pH values demonstrated that human peripheral blood lymphocytes regulate their internal pH, a process requiring expenditure of metabolic energy. In the pH range between 6.8 and 7.4, lymphocytes maintain a constant internal pH of 7.17 +/- 0.06 pH unit. Outside this range, intracellular pH changes with extracellular pH. The accuracy of this 19F pH probe has been confirmed by independent measurements of intracellular pH using equilibrium distributions of 5,5-dimethyloxazolidine-2,4-dione.

Cell Membrane Permeability↗
PubMed1982

Neurotransmitter metabolism in rat brain synaptosomes: effect of anoxia and pH.

Synaptosomes isolated from the rat cerebral cortex by means of a discontinuous Ficoll gradient carry out net, sodium-dependent, veratridine-sensitive accumulation of gamma-aminobutyric acid (GABA), serotonin, norepinephrine, and dopamine. The intrasynaptosomal contents of the four neurotransmitters are: 30.4 nmol/mg protein, 17.4 pmol/mg protein, 13.5 pmol/mg protein, and 21.2 pmol/mg protein, respectively. Anaerobic preincubation of synaptosomes causes an irreversible decrease in the rates of neurotransmitter accumulation but does not affect the rates of their release. The inhibitory effect of anaerobiosis is enhanced by increased concentration of [H+] (decreased pH) in the medium. The most sensitive is the uptake of dopamine, the least that of serotonin. The rates of neurotransmitter efflux are unaffected by anaerobiosis. Synaptosomes leak catecholamines, GABA, and serotonin into the medium when subjected to anaerobiosis, and reintroduction of oxygen is accompanied by a rapid reaccumulation of all four neurotransmitters. It is concluded that: (1) Responses of synaptosomes to anaerobiosis are remarkably similar to the behavior of intact brain in hypoxia and ischemia. (2) Neurotransmitter uptake systems are more sensitive to short periods of anaerobiosis than either the energy metabolism or ion transport. (3) Some neurotransmitter uptake systems are more easily damaged by anaerobiosis than others.

Aerobiosis↗
PubMed1982

The heat resistance of a ceramic identification device.

Identification of human remains by the teeth is a widely accepted forensic procedure. At present the principle dental technique used for identification of the dead is the post-mortem comparison method which involves a comparison of post-mortem dental findings and any available ante-mortem records. This study describes the results of an in vitro incineration investigation testing the heat resistance qualities of metallic intelligence data encoded on microchips comprising part of the Dentify system of identification. Detailed optical and scanning electron microscopic findings show that despite some alteration when exposed to high temperatures the metallic intelligence data itself remains clearly decipherable after exposure to a temperature of 1000 degrees C.

Ceramics↗
PubMed1982

Influence of presynaptic receptors on neuromuscular transmission in rat.

The presence and physiological significance of acetylcholine (ACh) receptors on motor nerve terminals was examined at the rat diaphragm neuromuscular junction. Intracellular recording techniques were used to monitor end-plate potentials (EPP), miniature end-plate potentials (MEPP), and resting potentials of the muscle fibers. Muscle action potentials were blocked by the cut-muscle technique. Quantal release was determined by the ratio EPP/MEPP, after correcting for nonlinear summation. Blockade of acetylcholinesterase with eserine and neostigmine was tested to determine the influence of residual ACh on transmitter release. Partial blockade of ACh receptors with curare was examined to further clarify the role of these presynaptic receptors. The experiments demonstrate that residual ACh inhibits transmitter release and that blockade of ACh receptors enhances transmitter release. It is concluded that presynaptic ACh receptors exist and that they serve an important physiological function. It is suggested that the presynaptic ACh receptors normally serve to limit transmitter release in a negative feedback pathway.

Animals↗
PubMed1982

Role of mitochondrial oxidative phosphorylation in regulation of coronary blood flow.

Regulation of coronary blood flow was studied in isolated rat hearts perfused under various metabolic conditions. Alterations in coronary flow were induced by hypoxia, amobarbital (Amytal) infusion, increase in work load of the heart, and adenosine infusion. Hypoxia induced, on the average, a 92.5% rise in coronary flow; 0.88 mM Amytal, a 85.7% increase; 12 microM adenosine, a 49.5% rise; and increased work load (elevation of the perfusion pressure from 6.9 kPa to 12.8 kPa), a 53.4% increase. In normoxia, adenosine, inosine, and hypoxanthine were present in the effluent in very low concentrations, and these greatly increased in response to hypoxia. In contrast, increased coronary flow caused by Amytal infusion or by elevated perfusion pressure was not accompanied by elevation in the effluent concentration of adenosine and its catabolites. Infusion of Amytal was followed by decrease in oxygen consumption of the heart and increase in oxygen tension in the effluent. This indicates that tissue oxygen tension per se can not be responsible for the regulation of coronary blood flow. Analysis of the data showed that under conditions in which there was a decrease in the tissue [ATP]free/[ADP]free[Pi] an increase in coronary flow was observed irrespective of the nature of the vasodilatory stimulus. It is concluded that mitochondrial oxidative phosphorylation provides a link between tissue oxygen metabolism and coronary blood flow. Mechanisms are discussed whereby changes in the cellular energy state ([ATP]free/[ADP]free[Pi]) are coupled to vasodilation, including possible direct effects on the vascular smooth muscle and/or generation of "second messengers."

Adenosine↗
PubMed1981

Surgical glove starch granuloma.

A variety of foreign bodies are capable of eliciting a granulomatous tissue response. Surgical glove lubricant powder is one source of foreign bodies. The consequences of talc and starch contamination of tissues are frequently reported for tissue sites outside the oral region. This article describes the clinical and histologic features of an intraoral starch granuloma.

Foreign Bodies↗
PubMed1981

Net uptake of gamma-aminobutyric acid by a high-affinity system of rat brain synaptosomes.

Rat brain synaptosomes isolated on discontinuous Ficoll gradient carry out rapid net uptake of gamma-aminobutyric acid through a high-affinity system (Km = 6.25 microM; Vmax = 1.2 nmol/min per mg of protein). The uptake of the labeled neurotransmitter is dependent on sodium concentration and is abolished by addition of 40 microM veratridine or 0.5 mM 2,4-diaminobutyric acid. Homoexchange in this preparation accounts for less than 10% of the measured uptake of gamma-amino[14C]butyric acid. It is concluded that the high-affinity transport exhibits properties characteristic of a system that is responsible for the rapid removal of gamma-aminobutyric acid from the synaptic cleft after neuronal transmission.

Aminobutyrates↗
PubMed1981

Effects of in vitro hypoxia and lowered pH on potassium fluxes and energy metabolism in rat brain synaptosomes.

Synaptosomes isolated on isosmotic Ficoll density gradients are an effective model for some aspects of neuronal function. They maintain metabolic energy levels ([ATP]/[ADP] [Pi]) and transplasma membrane electrical potentials very similar to those of neurons in the intact brain. The concentration of K+ in the external medium (K+-sensitive electrode), O2 uptake, and cytochrome c reduction (550 nm minus 540 nm) were simultaneously monitored in synaptosomal suspensions. Oxidative metabolism is the primary source of intrasynaptosomal ATP and at pH 7.4 anaerobiosis results in K+ leakage at 4.5 +/- 0.8 nmol/min/mg protein with glucose as substrate and 10.7 +/- 1.9 nmol/min/mg protein with lactate plus pyruvate (10:1) as substrate. Reintroduction of oxygen initiates complete (ouabain-sensitive) reuptake of K+ at initial rates of 35.4 +/- 3.2 nmol/min/mg protein and 18 +/- 1.7 nmol K+/min/mg protein, respectively. The rates of K+ leakage and reuptake fall when the pH is lowered from 7.4 to 6.0 but recover fully if the pH is raised to the original value. The rates of K+ release and uptake decrease when the Na+ concentration in the medium is decreased and increase when the Ca2+ concentration is decreased. The intrasynaptosomal [K+] under aerobic conditions was 77.3 +/- 3 mM and the calculated K+ diffusion potential was -72 mV. Anaerobic incubation of the synaptosomes from up to 20 min and at pH values from 7.4 to 6.0 did not produce irreversible impairment of any of the measured variables. These results suggest that permanent loss of brain function following prolonged hypoxia and ischemia is not due to irreversible damage to the synapses with respect to these parameters but rather to impairment of some other neuronal functions.

Adenosine Diphosphate↗
PubMed1981

Tympanic membrane perforations secondary to ventilation devices: special considerations.

Tympanic membrane perforations are uncommon following the use of ventilation devices; however, persistent perforations should be given special attention for squamous epithelial growth through the perforation to the undersurface of the tympanic membrane. Surgical management is indicated in these cases. Tympanic membrane grafting procedures are adequate for early involvement. Successful grafting depends on excision of the tympanic membrane involved with the undersurface growth of squamous epithelium. Mastoid extension of the squamous tissue ingrowth requires greater surgical exposure. Cases and their management are presented.

Adult↗
PubMed1980

Evaluation of enzyme systems and their regulation: the inapplicability of irreversible thermodynamics.

The formalism called irreversible thermodynamics has been examined for its applicability to the description of enzymic reactions. The basic assumption of this formalism is that the net flux through a reaction is related to the free energy change, a relationship which is usually assumed to be linear. These assumptions are shown to be approximately true for the trivial case of an enzyme reaction within 0.8 kJ/mol (0.2 kcal/mol) of equilibrium in the absence of changes in any regulatory parameters (such as inhibitor, activator, or enzyme concentration). For all other reaction conditions the net flux is not related to the free energy change and in special cases for which an apparent relationship is seen, it is not linear. Thus, application of the formalism of irreversible thermodynamics gives rise to qualitatively and quantitatively erroneous results and conclusions. Since most regulatory enzymic reactions are far from equilibrium, and the net reaction rate of such reactions is regulated by changes in inhibitors, activators and/or enzyme concentration, the formalism of irreversible thermodynamics is in general neither applicable nor useful in understanding the behavior of biological reaction systems.

Allosteric Regulation↗
PubMed1980

A specific uncoupler-binding protein in Tetrahymena pyriformis and Paracoccus denitrificans.

The uncoupler of mitochondrial oxidative phosphorylation, 2-nitro-4-azido-carbonylcyanide phenylhydrazone (N3CCP) which is capable of photoaffinity labeling has been used to examine the effect of uncouplers on the energy conserving membrane of Paracoccus denitrificans and Tetrahymena pyriformis. The N3CCP uncouples respiration in P. denitrificans and T. pyriformis cells with U1/2 values of 1.05 microM and 0.24 microM, respectively. Binding studies show the presence of 0.65 +/- 0.05 high affinity sites per cytochrome alpha with Kd of 0.5 +/- 0.1 microM in P. denitrificans membranes and 1.4 +/- 0.2 sites per cytochrome alpha 2 with a Kd of 0.4 +/- 0.1 microM in T. pyriformis membranes. Irradiation of [3H]-N3CCP bound to the membranes leads to a covalent linking of the radioactive uncoupler to a peptide of 10--15 kdaltons as analyzed by SDS-polyacrylamide gel electrophoresis. It is concluded that these two microbial systems contain a specific high affinity uncoupler binding site very similar to that of mammalian mitochondria (Katre, N.V. and Wilson, D.F. (1978) Arch. Biochem. Biophys. 191, 647--656).

Acetonitriles↗
PubMed1980

Interactions of cytochrome c with mitochondrial membranes. Binding to succinate-cytochrome c reductase.

Methyl-4-azidobenzoimidate was reacted with horse heart cytochrome c to give a photoaffinity-labeled derivative of this heme protein. The modified cytochrome c bound to cytochrome c-depleted mitochondria with the same Kd as native cytochrome c and restored oxygen uptake to the same extent. Irradiation of cytochrome c-depleted mitochondrial membranes with 3- to 4-fold excess of photoaffinity-labeled cytochrome c over cytochrome c oxidase resulted in covalent binding of the derivative to the membranes. Fractionation of the irradiated mitochondria in the presence of detergents and salts followed by chromatography on an agarose Bio-Gel-A-5m showed that the labeled cytochrome c was bound covalently to succinate-cytochrome c reductase. The covalently bound cytochrome c was active in mediating electron transfer between its reductase and oxidase. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of the succinate-cytochrome c reductase containing photoaffinity-labeled 125I-cytochrome c showed that the reductase contained a protein binding site for cytochrome c. It is suggested that cytochrome c1 is the most likely site for the cytochrome c binding in mitochondria in situ.

Affinity Labels↗
PubMed1980

Relationship of transmembrane pH and electrical gradients with respiration and adenosine 5'-triphosphate synthesis in mitochondria.

The mechanism of mitochondrial oxidative phosphorylation and its regulation have been studied by using suspensions of isolated rat liver mitochondria. Parallel measurements were made of mitochondrial volume, respiration, transmembrane pH and electrical gradients, and adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP), and inorganic phosphate (Pi) concentrations under various experimental conditions. The transmembrane electrical gradients were calculated from the equilibrium distributions of [3H]-triphenymethylphosphonium (TPMP+), [3H]tribenzylmethylammonium (TBMA+), and K+ (plus valinomycin). The transmembrane distributions of labeled acetate, methylamine, and 5,5-dimethyloxazolidine-2,4-dione were used for the calculation of pH gradients. Evaluation of the data shows that the respiratory rate is strictly correlated with [ATP]/([ADP][Pi]) (free energy of ATP synthesis), whereas there is no consistent correlation between the transmembrane electrical potential, the pH gradient, or the total "protonmotive force" (delta muH+) and the respiratory rate. Thermodynamic analysis indicates that, in order for the proton electrochemical gradient to serve as an intermediate in ATP synthesis, from three to seven H+ would have to be transported per each ATP synthesized, depending on the experimental conditions. These results suggest that the proton electrochemical gradient may not serve as a primary intermediate in oxidative phosphorylation.

Adenosine Diphosphate↗
PubMed1980

Regulation of cellular energy metabolism: the Crabtree effect.

The Crabtree effect (inhibition of respiration by glycolysis) is observed in cells with approximately equal glycolytic and respiratory capacities for ATP synthesis. Addition of glucose to aerobic suspensions of glucose-starved cells (Sarcoma 180 ascites tumor cells) causes a burst of respiration and lactate production due to ATP utilization for glucose phosphorylation by hexokinase and phosphofructokinase. This burst of activity is followed by inhibition of both respiration and glycolysis, the former to below the value before glucose addition (Crabtree effect). Both the respiratory rate and the glycolytic flux appear to be regulated by the cytosolic [ATP]/[ADP][Pi] albeit by completely different mechanisms. Respiration is regulated by the free energy of hydrolysis of ATP, such that the rate increases as the [ATP]/[ADP][Pi] decreases and decreases as the [ATP]/[adp][Pi] increases. The regulatory enzymes of glycolysis are activated by ADP (AMP) and Pi and inhibited by ATP. Thus both respiration and glycolysis increase or decrease as the [ATP]/[ADP][Pi] decreases or increases. The parallel regulation of both ATP-producing pathways by this common metabolite ratio is consistent with the cytoplasmic [ATP]/[ADP][Pi] being an important determinant of homeostatic regulation of cellular energy metabolism.

Adenosine Diphosphate↗