Biomedical subjects

D F Wilson

Publications and source records attributed to D F Wilson.

At least 163 records · Page 9Linked to original sources

PubMed1986

beta-2-Aminobicyclo-(2.2.1)-heptane-2-carboxylic acid. A new activator of glutaminase in intact rat liver mitochondria.

beta-(+/-)-2-Aminobicyclo-(2.2.1)-heptane-2-carboxylic acid (BCH) stimulated, in a concentration-dependent manner, the formation of glutamate by mitochondria isolated from rat liver and incubated with 20 mM glutamine. Maximum enhancement was seen with 10 mM BCH while 5 mM leucine was without effect. The initial lag in the rate of glutamate formation was not eliminated by BCH. Preincubation of the mitochondria without glutamine also did not abolish the lag period; to the contrary, it resulted in a progressive deactivation of the glutaminase. The decrease in enzyme activity during the preincubation without glutamine was partially reversed by the addition of either 10 mM BCH or 1.4 mM NH4Cl and was essentially abolished by their combined action. The apparently sigmoid rise in the activity of glutaminase with increasing concentration of glutamine became hyperbolic in the presence of 1.4 mM NH4Cl. BCH stimulated the NH4Cl-activated glutaminase in the entire range of glutamine concentrations studied (2-40 mM) without changing the S50 value. In mitochondria disrupted by repeated cycles of freezing and thawing, the enzymatic activity was maximal even in the absence of BCH. It is postulated that BCH is a potent activator of mitochondrial glutaminase and that manifestation of its action requires intact organelle structure. In addition, it is concluded that BCH-induced stimulation of glutamine catabolism in isolated hepatocytes (Zaleski, J., Wilson, D. F., and Erecinska, M. (1986) J. Biol. Chem. 261, 14082-14090) is the consequence of activation of the mitochondrial glutaminase.

Amino Acids↗

PubMed1986

The role of glial cells in regulation of neurotransmitter amino acids in the external environment. II. Mechanism of aspartate transport.

Active, high-affinity (Km = 4.4 microM) D-aspartate transport in C6 astrocytoma cells has been investigated. Uptake of radioactive D-aspartate was competitively inhibited by L-aspartate (Ki = 8.5 microM) and L-glutamate (Ki = 0.95 mM) and was essentially independent of pH between 6.2 and 7.8. The rate of uptake of labeled D-aspartate and its maximum accumulation ratio, [Asp]i/[Asp]e increased as the second power of the transmembrane electrical potential (measured by the potassium concentration gradient, [K+]i/[K+]e) which indicates that aspartate is transported with a net charge of +2. Aspartate transport rate and gradient also increased as the second power of the sodium concentration gradient, [Na+]e/[Na+]i, indicating that two Na+ are transported inward with each aspartate. The maximum gradient measured from total intra- and extracellular concentrations of aspartate showed the same dependence on electrical potential and sodium concentration gradient as that determined from the distribution of [3H]D-aspartate. This indicates that energy for aspartate uptake is provided by a combination of transmembrane electrical potential and sodium concentration gradient. At physiological [Na+]e (140 mM) and [K+]e (3.5-5 mM) the energy available for aspartate uptake substantially exceeded the maximum aspartate gradient. It is suggested that aspartate uptake by C6 cells is kinetically prevented from attaining high concentration differences and that the excess of driving forces over accumulation ratio ensures that glial high-affinity transport systems for amino acid neurotransmitters function in vivo predominantly in the direction of net uptake.

Animals↗

PubMed1986

The oxygen dependence of cellular energy metabolism.

In isolated perfused rat hearts coronary flow appears to be regulated through the tissue energy level ([ATP]free/[ADP]free[Pi]), a metabolic product of oxygen metabolism by mitochondria, and not by the tissue oxygen tension. Mitochondrial oxidative phosphorylation in intact cells and tissues is dependent on oxygen tension throughout the physiological range. This allows oxidative phosphorylation to serve as an important tissue oxygen sensor. The apparent Km of mitochondrial cytochrome c oxidase for oxygen is dependent on the pH of the suspending medium, [ATP]/[ADP][Pi] and the state of reduction of cytochrome c. When physiological levels of [ATP]/[ADP][Pi], pH and cytochrome c reduction are used, oxidative phosphorylation by suspensions of isolated mitochondria has an oxygen dependence similar to that observed in intact cells.

Animals↗

PubMed1986

A new method for measuring oxygen concentration in biological systems.

The oxygen dependent quenching of phosphorescence, if measured by the oxygen dependence of the phosphorescence lifetime, can be used as a sensitive and precise measure of oxygen concentration. The result is an optical method for measuring oxygen which is easily calibrated and the calibration may, depending on the experimental conditions, be stable for months to years. Although much remains to be done to realize the full potential of this method of measuring oxygen, it has great promise and we are proceeding with its development.

Kinetics↗

PubMed1986

Esophageal adenocarcinoma metastatic to the maxilla.

Although metastatic malignant lesions in the maxilla are exceedingly rare, case reports of such lesions do appear occasionally in the literature. This article describes the clinical and pathologic features of an esophageal adenocarcinoma metastatic to the maxilla. The initial maxillary appearance of the tumor mimicked primary dental pathosis and, in addition, was the presenting sign of malignant disease in the patient. The relevant literature is reviewed, and the processes by which metastases form are discussed.

Adenocarcinoma↗

PubMed1986

Transport of cysteate by synaptosomes isolated from rat brain: evidence that it utilizes the same transporter as aspartate, glutamate, and cysteine sulfinate.

Synaptosomes isolated from rat brain accumulated cysteic acid by a high-affinity transport system (Km = 12.3 +/- 2.1 microM; Vmax = 2.5 nmol mg protein-1 min-1). This uptake was competitively inhibited by aspartate (Ki = 13.3 +/- 1.8 microM) and cysteine sulfinate (Ki = 13.3 +/- 2.3 microM). Addition of extrasynaptosomal cysteate, aspartate, or cysteine sulfinate to synaptosomes loaded with [35S]cysteate induced rapid efflux of the cysteate. This efflux occurred via stoichiometric exchange of amino acids with half-maximal rates at 5.0 +/- 1.1 microM aspartate or 8.0 +/- 1.3 microM cysteine sulfinate. Conversely, added extrasynaptosomal cysteate exchanged for endogenous aspartate and glutamate with half-maximal rates at 5.0 +/- 0.4 microM cysteate. In the steady state after maximal accumulation of cysteate, the intrasynaptosomal cysteate concentrations exceeded the extrasynaptosomal concentrations by up to 10,000-fold. The measured concentration ratios were the same, within experimental error, as those for aspartate and glutamate. Depolarization, with either high [K+] or veratridine, of the plasma membranes of synaptosomes loaded with cysteate caused parallel release of cysteate, aspartate, and glutamate. It is concluded that neurons transport cysteate, cysteine sulfinate, aspartate, and glutamate with the same transport system. This transport system catalyzes homoexchange and heteroexchange as well as net uptake and release of all these amino acids.

Amino Acids, Sulfur↗

PubMed1986

A role for transglutaminase in neurotransmitter release by rat brain synaptosomes.

Rat brain synaptosomes exhibit calcium-dependent transglutaminase activity. This activity, measured in detergent-treated or sonicated preparations, was six- to sevenfold lower than that in the liver. The synaptosomal transglutaminase was inhibited by various amines and alpha-difluoromethylornithine, compounds known to inhibit activity of this enzyme in other tissues. The inhibitors of transglutaminase induced release of catecholamines, but not of gamma-aminobutyric acid, from synaptosomes both under basal and K+-stimulated conditions. The concentrations of the agents that caused stimulation of catecholamine release were approximately the same as those that inhibited the activity of transglutaminase. Stimulation of release was largely reduced by the withdrawal of calcium from the incubation medium. Inhibitors of transglutaminase had little effect either on the uptakes of neurotransmitters or the amounts of deaminated products of catecholamine degradation released into the medium. It is suggested that a synaptosomal transglutaminase is involved in suppressing vesicular release of catecholamines by resting (nondepolarized) neurons and that this action may also be a part of negative feedback control which prevents excessive transmitter release at the synapse during increased neuronal activity.

Animals↗

PubMed1986

Chronic effects of paraoxon on transmitter release and the synaptic contribution to tolerance.

The chronic effects of sublethal injections of the cholinesterase inhibitor, paraoxon, on transmitter release were examined in the rat. Rats were chronically treated daily with the organophosphate, paraoxon (0.3 mg/kg b.wt.), that is known to inhibit acetylcholinesterase irreversibly. Severe symptoms of intoxication were evident in animals during the first few days of treatment but by day 11 the symptoms disappeared despite continued treatment and depression in acetylcholinesterase. Intracellular recording techniques were used to determine if the observed behavioral changes to chronic treatment could be correlated with changes in neuromuscular transmission. Measurements of MEPPs demonstrated that tolerance could not be attributed to a decrease in postsynaptic sensitivity. Measurements of EPPs, quantal release and binomial statistical parameters demonstrated that tolerance can be correlated with presynaptic changes. Chronic treatment with paraoxon decreased transmitter release and this can be attributed to a decrease in the transmitter store and mobilization ability. It is suggested that the depression in quantal release at the neuromuscular junction and at cholinergic synapses could account for behavioral tolerance.

Acetylcholine↗

PubMed1985

Kainate-induced uptake of calcium by synaptosomes from rat brain.

Kainic acid induces a rapid increase in 45Ca2+ uptake by crude synaptosomal fractions isolated from rat brain. This enhanced Ca2+ permeability occurs with a half-time of approx. 1 s, similar to the fast phase of depolarization-induced calcium uptake. The depolarization-induced uptake of calcium is inhibited 85% by 3 mM CoCl2, 80% by 100 microM quinacrine and 50% by 15 microM trifluoperazine while these agents had little effect on the kainate-induced uptake. It is proposed that kainate induces receptor-mediated opening of a class of calcium channels with properties different from those of the voltage-dependent channels.

Animals↗

PubMed1985

Activation of tyrosine hydroxylase in the central nervous system by anaerobiosis.

Subjecting either P2 fraction or purified synaptosomes isolated from rat brain to periods of anoxic incubation at 30 degrees resulted in activation of dopamine synthesis from tyrosine. This activation was approximately 2.5-fold when the anoxic incubation was carried out at pH 6.2 but was not significant when the pH was 7.4. Measurements of the tyrosine hydroxylase activity at pH 6.2 in Triton X-100-treated preparations of the P2 fraction showed that, after 20 min of anaerobiosis, the Km for pterine cofactor decreased by 39% and the Ki for dopamine increased by 44%; there was no change in the Km for tyrosine. Half-maximal activation of dopamine synthesis occurred in 10 min of anaerobic incubation, and the reversal upon addition of oxygen had a half-time of 15 min. Addition of forskolin or dibutyryl cyclic AMP to anaerobic incubations of P2 fraction did not result in significant activation of dopamine synthesis. Either the removal of calcium or the addition of calmodulin inhibitor, trifluoperazine, substantially decreased the activation of dopamine synthesis induced by periods of anaerobiosis. It appears that during anoxic incubation tyrosine hydroxylase underwent an activation which occurred over a period of minutes, was stable to detergent treatment, and was fully reversed over a period of minutes following reoxygenation. This activation was, at least in part, dependent on the presence of calcium and was sensitive to the calmodulin antagonist trifluoperazine.

Anaerobiosis↗

PubMed1985

The effect of glucagon and adrenergic agonists on intracellular pH of isolated rat hepatocytes.

19F NMR was used to measure intracellular [H+] of hepatocytes before and after incubation with glucagon and adrenergic agonists at their concentrations which give maximal stimulation of both glucose and urea production. Intracellular and extracellular pH was determined from the chemical shifts in resonances of alpha-difluoromethylalanine. The alterations in intracellular [H+] with agonist treatment were, in all cases, found to be less than 0.1 pH unit in the pH range 6.7-7.8. It is concluded that changes in concentration of the intracellular [H+] do not play a significant role in the stimulation of urea and glucose production caused by these hormonal effectors.

Animals↗

PubMed1985

Effects of ATP gamma S in isolated rat brain synaptosomes.

The effects of ATP gamma S, a slowly hydrolyzable analogue of ATP, were investigated in the preparation of synaptosomes isolated from rat cerebral cortex. It was found that addition of [35S]ATP gamma S resulted in substantial magnesium-dependent incorporation of 35S into synaptosomal proteins which was prevented completely by ATP. The most prominently labeled polypeptides were those with apparent molecular weights of 100,000; 84,000; 74,000; 62,000; 55,000; 48,000; and 41,000. The rate and extent of thiophosphorylation were unaffected by addition of cAMP, veratridine or sodium fluoride. ATP gamma S at 50-100 microM had no effect on either uptake or release of gamma-aminobutyric acid (GABA) and dopamine; at a concentration of 1 mM it inhibited incorporation of dopamine by about 20%. This inhibition was also seen with 1 mM GTP, beta, gamma-methylene-adenosine 5'-triphosphate and adenylylimidodiphosphate, which suggests that the nucleotide triphosphates themselves, and not membrane protein phosphorylation, were responsible for the effect observed. It is concluded that ATP gamma S is an effective tool for studying the possible role of ATP released in synaptic transmission. The results obtained thus far suggest that neither extrasynaptosomal ATP nor phosphorylation of external proteins of the presynaptic membrane is sufficient for modulation of neurotransmitter uptake or release. They may, however, play a role in combination with other conditions.

Adenosine Diphosphate↗

PubMed1985

The distribution of inorganic phosphate and malate between intra- and extramitochondrial spaces. Relationship with the transmembrane pH difference.

The steady-state distribution of inorganic phosphate and malate between the intra- and extramitochondrial spaces was measured in suspensions of nonrespiring and respiring rat liver mitochondria in which the transmembrane pH difference was incrementally varied. In respiration-inhibited mitochondria, the slope of log [Pi]in/[Pi]out (ordinate) versus delta pH approached 2 by either chemical or isotopic determination of [Pi], and the slope of log [malate]in/[malate]out versus delta pH was 2.0 with an extrapolated log [Pi]in/[Pi]out value of 0.3 at delta pH = 0. We conclude that the distribution of Pi and malate for nonrespiring mitochondria were quantitatively consistent with those predicted by exchange of Pi- for OH- (or cotransport with H+) and of malate 2- for Pi2-. In respiring mitochondria using glutamate + malate as substrate, there was very little pH dependence of Pi or malate accumulation (the slopes were less than 0.5) unless n-butylmalonate (inhibitor of Pi-dicarboxylate exchange) was added before the glutamate and malate, in which case the distribution patterns at delta pH less than 0.4 were similar to those in nonrespiring mitochondria. In either case, however, after reaching a maximal value of 1.1, log [Pi]in/[Pi]out did not further increase with increasing delta pH. Thus, in normally metabolizing mitochondria, the distributions of Pi and malate are not directly correlated with the difference in pH across the membrane.

Animals↗

PubMed1985

pH dependence of hormonal regulation of gluconeogenesis and urea synthesis from glutamine in suspensions of hepatocytes.

Hepatocytes from rats deprived of food for 48 h synthesized glucose and urea from glutamine at a rate which, at pH 7.3, was markedly stimulated (175-250%) by dibutyryl cAMP, phenylephrine, and norepinephrine, in agreement with previous investigators. These effectors also stimulated respiration, elevating ATP production by the amount required for the increase in glucose and urea synthesis. Both the basal and stimulated rates were strongly pH dependent with maxima in the region of pH 7.2-7.6 (urea synthesis) and 7.2-7.5 (glucose synthesis) and declined rapidly on either side of these pH values. The inhibitions at acid and alkaline pH were neither due to lack of energy nor to limitation in glutamine uptake. The intracellular concentrations of aspartate, glutamate, and glutamine were lower at pH 6.7 than at pH 7.3 and were differently affected by dibutyryl cAMP and phenylephrine at the two pH values investigated. When calcium was omitted from the suspending medium, the basal rates of glucose and urea production were decreased as was stimulation by the effectors, phenylephrine completely, and the others partially. The stimulations by phenylephrine and dibutyryl cAMP were additive under all conditions tested. The pattern of metabolite changes indicates that although both effectors stimulated glutaminase and increased supply of aspartate to the argininosuccinate synthetase, dibutyryl cAMP gave greater activation of glutaminase whereas the adrenergic agonists gave greater stimulation of later steps on the biosynthetic pathways. It may be physiologically important than at acid pH both ureagenesis and gluconeogenesis are severely suppressed and cannot be effectively stimulated by the major hormonal regulators of these pathways.

Adenosine Diphosphate↗

PubMed1985

Inherited craniofacial fibrous dysplasia.

Craniofacial fibrous dysplasia occurring in a mother and two of her three daughters is reported. In all three patients, the condition was characterized by extensive bilateral and bimaxillary involvement. All patients exhibited radiographic evidence of abnormal maxillary and mandibular bone and tooth displacement. Morphologic examination of deciduous teeth from the daughters showed replacement of normal pulp tissue by fibro-osseous lesion. Because of the familial occurrence of the cases, it is proposed that they comprise an example of inherited craniofacial fibrous dysplasia.

Adult↗

PubMed1985

The effect of cholinergic agonists on coronary flow rate and oxygen consumption in isolated perfused rat heart.

The metabolic and circulatory consequences of activation of the muscarinic receptor(s) were investigated by local administration of acetylcholine and its three analogues (bethanechol, carbachol and methacholine) into beating and KCl-arrested perfused rat hearts. Acetylcholine and the three other choline esters caused vasoconstriction in both types of preparations and this vasoconstriction was accompanied by a decrease in oxygen consumption. In most cases the dose-response curves were biphasic and changes in coronary flow paralleled those in oxygen consumption. Both phenomena were abolished by administration of atropine and either removal of calcium or infusion of verapamil but were unaffected by addition of the adrenergic alpha-blocker, prazosin, and the adrenergic beta-blocker, propranolol. Infusions of low concentrations of the cholinergic agonists were accompanied by increases in the myocardial phosphorylation state ratio [( ATP]free/[ADP]free[Pi]) which correlated with the simultaneous decreases in oxygen consumption and coronary flow. It is suggested that muscarinic receptors responsible for vasoconstriction in perfused rat heart are located not only on coronary vessels but also on the cardiac muscle cells. Activation of the former receptors induces vasoconstriction by direct action on the vascular smooth muscle while activation of the latter receptors induces vasoconstriction indirectly by decreasing cardiac work and increasing the myocardial [ATP]free/[ADP]free[Pi] ratio. The results also show that stimulation of muscarinic receptor(s) and the consequent metabolic and vasoregulatory responses are coupled to calcium movements.

Animals↗

PubMed1985

Some effects of ischaemia and hyperglycaemia on neurotransmitter metabolism in rat brain.

30 min of low flow cerebral ischaemia (0.12 ml/min) was imposed on normal and hyperglycaemic rats. Some were allowed to recover for up to 10 h. Whole tissue homogenates, or synaptosomes from the brains of each group were examined for content of noradrenaline, dopamine and 5-HT of defined anatomical regions. In normoglycaemic rats immediately after ischaemia there was a significant reduction in the content of all three neurotransmitters in cortex, striatum and hippocampus but not in diencephalon (except possibly 5H-T) or brain stem. The amounts of all three neurotransmitters returned to control values after about 30 min reperfusion and remained so for 10 h. By contrast, no changes in neurotransmitter levels were found either during or after ischaemia in hyperglycaemic rats. Dopamine synthesis rates in the striatum were increased after ischaemia and returned to control levels in 2-3 h in normoglycaemic rats but were unaltered in hyperglycaemic animals. Binding sites for spiroperidol and LSD were compared in cortical synaptosomes and whole cortex homogenates in the presence and absence of agonists and antagonists. Both types of membrane showed similar binding properties and there was no significant change in number or character of binding sites immediately following ischaemia or during 10 h of brain reperfusion. The significance of these findings is discussed in relation to ischaemic brain damage.

Animals↗

PubMed1985

Histopathology of the mandibular condyle.

Histological sections of mandibular condyles surgically removed from 40 female and 12 male patients who failed to respond to conservative management of temporomandibular joint related problems were examined. The proliferation, intermediate and cartilage zones were frequently either partly or completely absent in older subjects confirming previously held views of the functional significance of these zones. There was no obvious sex differences in the histological appearance of the condyle and evidence of inflammatory change was rarely seen.

Adolescent↗