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D E Atkinson

Publications and source records attributed to D E Atkinson.

At least 19 recordsLinked to original sources
PubMed2005

MDR1 P-gp expression and activity in intact human placental tissue; upregulation by retroviral transduction.

This study investigates P-gp activity in placental villous fragments and the possibility of upregulating its expression and function by retroviral transduction. In fresh fragments, cyclosporin A caused a significant increase in 3H-vinblastine accumulation (187+/-48% at 180 min n=4), consistent with multi-drug resistance activity. After 7 days in culture, villous fragments showed a similar increase in 3H-vinblastine accumulation (143+/-10% at 180 min n=4), which was not significantly different from that in fresh tissue. Following transduction, immunohistochemistry revealed increased P-gp expression. However, the distribution of the protein differed from that in controls, with P-gp being located throughout the tissue as opposed to the normal specific location on the maternal facing plasma membrane. Transduced explants showed a significantly larger increase in 3H-vinblastine accumulation in the presence of cyclosporin A than control explants (245+/-15.5% at 180 min, n=4), suggesting reduced capacity to efflux vinblastine. This study demonstrates P-gp activity in intact placental tissue which is maintained in explant culture. Retroviral transduction of P-gp to such tissue leads to increased but undirected expression of the protein. The consequent increased activity at sites such as the basal, fetal facing, plasma membrane probably explains the increased substrate accumulation within the tissue.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1998

Diagnosis of heterotaxy syndrome by fetal echocardiography.

We report a series of 13 consecutive patients diagnosed in utero with heterotaxy syndrome in which we found a slight predominance (8 of 13) of fetuses with right atrial isomerism. In previous studies in which diagnosis of left or right isomerism was made based upon findings at fetal echocardiography, there has been a preponderance of infants (95%) diagnosed with left isomerism.

Echocardiography, Doppler↗
PubMed1995

pH heterogeneity in aged hypertensive rat hearts distinguishes reperfused from persistently ischemic myocardium.

Myocardium reperfused following ischemia may contain regions that are adequately perfused but temporarily dysfunctional or metabolically abnormal, and regions that are persistently ischemic. By examining heterogeneity in the inorganic phosphate (Pi) resonance in 31P NMR spectra obtained for isolated perfused young v old hypertensive rat hearts made globally ischemic and then reperfused, areas of persistent intracellular acidosis (pHi about 6) were distinguished from recovering tissue (pHi about 7). The extent of persistent regional acidosis reported by Pi heterogeneity in the 31P NMR spectrum of the reperfused heart correlates well with (1) whole heart flow deficit measured as coronary flow, (2) the decrease in extracellular (primarily vascular) 23Na NMR resonance area measured using 23Na NMR and shift reagent, and (3) regional flow deficits identified using perfusate-borne dye markers. Persistent intracellular acidosis during reperfusion was observed only in the hearts of aged hypertensive rats, confirming that the vasculature of these animals is more susceptible to ischemic injury than both younger hypertensive rats and age-matched controls. The portion of the heart that is reperfused following only 16 min of global ischemia is metabolically abnormal, showing incomplete recovery of ATP and pH.

Adenosine Triphosphate↗
PubMed1992

Gestational changes in Ca2+ transport across rat placenta and mRNA for calbindin9K and Ca(2+)-ATPase.

The unidirectional maternofetal clearance (Kmf) of 45Ca was measured across the rat placenta over the last one-third of gestation. Kmf for 45Ca normalized to its diffusion coefficient in water (Kmf/Dw) increased 72-fold between days 15 and 22 of gestation from 3.5 +/- 0.3 to 253.1 +/- 22.0 cm/g placenta, respectively. At 15 and 18 days of gestation, Kmf/Dw for 45Ca was similar to Kmf/Dw for the paracellular marker [14C]mannitol, but at 21 and 22 days of gestation, Kmf/Dw for 45Ca was significantly higher than Kmf/Dw for [14C]mannitol, indicating that an additional route of transfer, other than diffusion, becomes available to calcium during this period. Northern hybridization analysis demonstrated that rat placental calbindin9K-to-beta-actin mRNA ratio increased 135-fold between 15 and 22 days of gestation and was temporally associated with the gestational increase in Kmf/Dw for 45Ca. In contrast, rat placental Ca(2+)-ATPase-to-beta-actin mRNA ratio increased only two- to threefold over the same gestational period and did not mirror the gestational changes in calcium clearance. These trends suggest that the expression of placental calbindin9K, but not Ca(2+)-ATPase, may be rate limiting to placental calcium transport in the rat.

Actins↗
PubMed1991

Analysis of 23Na NMR spectra from isolated perfused hearts.

The 23Na NMR spectra obtained from isolated hearts perfused with buffer containing the paramagnetic shift reagent dysprosium triethylenetetraminehexaacetic acid, Dy(TTHA)3-, are complex and contain a number of overlapping peaks of different intensities. Spectra from rat, rabbit, guinea pig, and ferret hearts obtained during periods of control perfusion are similar and undergo similar changes when the hearts are subjected to periods of ischemia and reflow. The contributions from the intracellular, interstitial, vascular, and bath compartments to the multiple peaks in the spectra of rats hearts have been assigned. The significant contributions to these spectra of bulk magnetic susceptibility effects and incomplete mixing have been demonstrated through a series of modeling experiments. Since the spectra from hearts of different species are so similar, the peak assignments made for the rat are applicable to spectra from rabbit, guinea pig, and ferret hearts as well. This work provides a framework for quantitative analysis of the spectral changes which occur during conditions such as ischemia and reflow.

Animals↗
PubMed1991

Development of passive permeability characteristics of rat placenta during the last third of gestation.

To investigate how the passive permeability of the rat placenta changes during the last third of gestation we measured the unidirectional maternofetal clearance (Kmf) of three polar nonelectrolytes. We found that Kmf (microliters.min-1.g placenta-1) for all three tracers increased between days 15 and 22 (term is 23 days). The increase was less for the largest tracer, [14C]inulin (3.5-fold increase from 0.5 +/- 0.1 to 1.6 +/- 0.1 microliters.min-1.g placenta-1), than for the smaller tracers, [14C]mannitol (7-fold increase from 1.6 +/- 0.1 to 10.9 +/- 0.4 microliters.min-1.g placenta-1) and 51Cr-labeled EDTA (9.5-fold increase from 0.8 +/- 0.04 to 7.6 +/- 0.7 microliters.min-1.g placenta-1). These data suggest that whilst the overall permeability of the rat placenta increases during this period of pregnancy, restriction to the diffusion of large (greater than or equal to 1.5 nm molecular radius) polar nonelectrolytes also increases.

Animals↗
PubMed1990

Effect of level of dietary protein on arginine-stimulated citrulline synthesis. Correlation with mitochondrial N-acetylglutamate concentrations.

Increases in dietary protein have been reported to increase the rate of citrulline synthesis and the level of N-acetylglutamate in liver. We have confirmed this effect of diet on citrulline synthesis in rat liver mitochondria and show parallel increases in N-acetylglutamate concentration. The magnitude of the effect of arginine in the suspending medium on citrulline synthesis was also dependent on dietary protein content. Mitochondria from rats fed on a protein-free diet initially contained low levels of N-acetylglutamate, and addition of arginine increased the rate of its synthesis. Citrulline synthesis and acetylglutamate content in these mitochondria increased more than 5-fold when 1 mM-arginine was added. A diet high in protein results in mitochondria with increased N-acetylglutamate and a high rate of citrulline synthesis; 1 mM-arginine increased citrulline synthesis in such mitochondria by only 36%. The concentration of arginine in portal blood was 47 microM in rats fed on a diet lacking protein, and 182 microM in rats fed on a diet containing 60% protein, suggesting that arginine may be a regulatory signal to the liver concerning the dietary protein intake. The rates of citrulline synthesis were proportional to the mitochondrial content of acetylglutamate in mitochondria obtained from rats fed on diets containing 0, 24, or 60% protein, whether incubated in the absence or presence of arginine. Although the effector concentrations are higher than the Ka for the enzymes, these results support the view that concentrations of both arginine and acetylglutamate are important in the regulation of synthesis of citrulline and urea. Additionally, the effects of dietary protein level (and of arginine) are exerted in large part by way of modulation of the concentration of acetylglutamate.

Animals↗
PubMed1990

Response of rat liver glutaminase to pH, ammonium, and citrate. Possible regulatory role of glutaminase in ureagenesis.

Liver glutaminase is stimulated by an increase in NH4+ concentration and NH4+ is an absolute requirement for activity at approximate physiological concentrations of phosphate and glutamine. Increases in the concentration of NH4+ cannot, however, overcome the inhibitory effect of a decrease in pH. In addition, the concentration of NH4+ required for half-maximal rate decreases as pH increases. This decrease is the result of two factors: a direct effect of pH on the apparent affinity of the enzyme for NH4+, and an indirect effect of pH brought about by an increase in the apparent affinity of the enzyme for phosphate which results in a further decrease in the M0.5 for NH4+. In addition, liver glutaminase responds strongly to the concentration of citrate over a physiologically relevant range at approximate physiological concentrations of NH4+, phosphate, and glutamine. An increase in citrate concentration stimulates glutaminase by increasing the affinity of the enzyme for glutamine. The apparent affinity of the enzyme for citrate increases as pH increases. The strong response of liver glutaminase to pH, NH4+, and citrate and the fact that the hydrolysis of glutamine can supply metabolites and effectors for urea synthesis suggest a possible regulatory role of glutaminase in ureagenesis.

Ammonia↗
PubMed1990

Response of rat liver glutaminase to magnesium ion.

The activity of rat liver glutaminase from sedimented fractions of freeze-thawed mitochondria is strongly affected by variation in the Mg2+ concentration within the approximate physiological range of activators. A rise in the Mg2+ concentration stimulates glutaminase by increasing the apparent affinity of the enzyme for its positive modifier phosphate. With the addition of 4 mM Mg2+ the M0.5 for phosphate activation decreased from 18 to 9.5 mM at pH 7.1, 10 to 5.8 mM at pH 7.4 and 6.4 to 4.0 mM at pH 7.7. The result is an increase in the apparent affinity of the enzyme for glutamine. With the addition of 4 mM Mg2+ the S0.5 of glutaminase for glutamine decreased from 24 to 13 mM at pH 7.1, 14 to 9.6 mM at pH 7.4, and remained unchanged at 8.2 mM at pH 7.7. Since Mg2+ stimulates glutaminase, as does a rise in pH (Szweda, L.I. and Atkinson, D.E. (1989) J. Biol. Chem. 264, 15357-15360), by increasing the apparent affinity of the enzyme for phosphate, it reduces the inhibitory effect of a decrease in pH and/or phosphate concentration over a physiologically relevant range.

Animals↗
PubMed1990

Effects of ultrasound on placental transfer during the last third of gestation in the rat.

The effect of ultrasound on placental transfer in the rat has been measured in vivo. At 15, 18, 21 and 22 days of gestation one uterine horn of each anaesthetized rat was exposed, after laparatomy, to ultrasound. The control horn was effectively shielded. Radio-labelled tracers were then injected into the maternal circulation via a jugular vein cannula and fetuses removed simultaneously from both experimental and control horns at times predetermined to ensure minimal backflux from fetus to mother. Samples of maternal plasma were also taken at suitable intervals. The radioactive content of fetuses and plasma were measured and a unidirectional materno-fetal clearance value (Kmf) for each fetus calculated. It was shown that in rats whose body temperature was accurately maintained at 37 degrees C, ultrasound at the level used in the present study had no effect on placental clearance. However, in rats whose body temperature was allowed to fall to 32 degrees C, a 3 min exposure to 1.1 MHz continuous wave ultrasound at 1 W cm-2 caused a significant (p less than 0.001) decrease in the placental clearance of 22Na (by 55%) and 51Cr-EDTA (by 45%) in rats at 15 days of gestation. These effects showed no dose response and were not observed in rats at any of the other stages of gestation which we investigated. However, in rats at 21 days of gestation the clearance of 45Ca2+ was significantly (p less than 0.01) reduced (by 30%) by a similar exposure to ultrasound. It is concluded that although an interesting biological effect of ultrasound at low body temperatures has been observed, it has little relevance for the normal use of ultrasound in obstetrics.

Animals↗
PubMed1990

Energetic correlates of cardiac failure: changes in the creatine kinase system in the failing myocardium.

To address the hypothesis that impaired ATP synthesis rates caused by changes in the creatine kinase system is an important mechanism underlying cardiac failure, we measured total creatine kinase activity, isoenzyme composition and creatine content in two animal models of hypertrophy with cardiac dysfunction, the spontaneously hypertensive rat in the transition to failure and the creatine-depleted hyperthyroid rat heart challenged by hypoxia. During the transition from stable compensated hypertrophy to failure characterized by decreased functional capacity, we found that total creatine kinase activity and particularly mitochondrial creatine kinase activity decreased. The decrease in functional capacity, the further increase in heart size and the derangements in the creatine kinase system did not occur if these animals were treated for 6 months with the antihypertensive agents, guanethidine or hydralazine. These results suggest that changes in the creatine kinase system occur coordinately with the transition to failure. To assess whether the changes in the creatine system may be causally linked to decreased functional capacity, we used 31P NMR spectroscopy of isolated perfused hearts to define the high energy phosphate content and cardiac performance of creatine-depleted (approximately 50%) hypertrophied hearts challenged by hypoxia. These hearts displayed greater susceptibility to hypoxic injury with regard to both systolic and diastolic function during and following hypoxia. We also measured total creatine kinase activity in right ventricular biopsy specimens from patients with various forms of cardiomyopathy and low ejection fractions, and found a positive correlation between total creatine kinase activity and ejection fraction. Taken together, these results support the hypothesis that decreasing the energy reserve for ATP synthesis renders the heart more susceptible to systolic and diastolic failure.

Animals↗
PubMed1989

Response of rat liver glutaminase to pH. Mediation by phosphate and ammonium ions.

The activity of rat liver glutaminase from sedimented fractions of freeze-thawed mitochondria is strongly affected by variation in pH over a physiologically relevant range at approximate physiological concentrations of activators. As pH increases from 7.1 to 7.7 at 0.7 mM ammonium and 10 mM phosphate, the S0.5 for glutamine decreases 3.5-fold, from 38 to 11 mM. This results in an 8-fold increase in reaction velocity at 10 mM glutamine. In addition, the M0.5 for phosphate activation decreases from 21 to 8.9 mM as pH increases from 7.1 to 7.7. This apparent effect of pH on the affinity of glutaminase for phosphate is similar to previous reports of the pH effect on activation by ammonium (Verhoeven, A. J., Van Iwaarden, J. F., Joseph, S. K., and Meijer, A. J. (1983) Eur. J. Biochem. 133, 241-244; McGivan, J. D., and Bradford, N. M. (1983) Biochim. Biophys. Acta 159, 296-302). Glutaminase does not respond to variation in pH between 7.1 and 7.7 when phosphate and ammonium are saturating. The effects of the two modifiers are additive. Each is still effective, as is pH, when the other is saturating. Therefore, it appears that the effects of pH on the apparent affinity of the enzyme for ammonium and phosphate account for the enzyme's response to pH. These results may help explain previous reports of minimal effects of pH on glutaminase at saturating concentrations of related substances (McGivan, J. D., Lacey, J. H., and Joseph, K. (1980) Biochim. J. 192, 537-542; Horowitz, M. L., and Knox, W. E. (1968) Enzymol. Biol. Clin. 9, 241-255; McGivan, J. D., and Bradford, N. M. (1983) Biochim. Biophys. Acta 759, 296-302). Glutaminase binds glutamine cooperatively with Hill coefficients ranging from 1.7 to 2.2, which suggests at least two and probably three or more interacting binding sites for glutamine. The strong response of liver glutaminase to pH and the fact that the reaction can supply metabolites for urea synthesis suggest a possible regulatory role of glutaminase in ureagenesis.

Ammonia↗
PubMed1989

Amiloride reduces maternofetal clearance of sodium across the near-term rat placenta.

The effect of amiloride on unidirectional maternofetal clearance across the perfused placenta has been studied in rats at 21 days gestation. A bolus dose of 1 mg kg-1 followed by an infusion of 0.6 mg h-1 into the maternal circulation caused a significant (P less than 0.02) decrease in 22Na clearance by 8%, whilst having no effect on the clearance of 51Cr-EDTA; a ten times higher dose of amiloride (10 mg kg-1, 6 mg h-1) did not reduce 22Na clearance further. Similar administration of an equivalent volume of saline to control animals showed no effect on either Na+ or Cr-EDTA clearances.

Amiloride↗
PubMed1987

Metabolic aspects of the regulation of systemic pH.

Catabolism of protein produces CO2, NH4+, and HCO3-. Mammals readily lose CO2 through the lungs, but the bicarbonate produced in metabolism of a typical diet (in humans, approximately 1 mol/day from approximately 100 g of protein) would cause alkalosis if not disposed of. Air-breathing animals solve this problem by incorporating NH4+ into organic compounds in which N is not protonated; thus each NH4+ ion loses a proton in the course of the synthesis. These protons serve to titrate HCO3-. In mammals, ureagenesis is the pathway by which protons are liberated from NH4+. The rate of ureagenesis therefore determines the rate of disposal of bicarbonate, and must be an important factor in the maintenance of pH homeostasis. Ammonium ion that is not needed for urea synthesis is packaged into glutamine by the liver. Hepatic glutamine synthetase is localized in the last rank of cells around the pericentral venule; thus glutamine synthetase cannot compete for NH4+ or interfere with the control of pH by urea synthesis. Ammonium excretion in the urine does not represent excretion of acid, and is not stoichiometrically related to renal generation of bicarbonate. The quantitatively major processes by which the HCO3-/CO2 ratio, and hence the pH, is regulated in blood and interstitial fluid are excretion of CO2 through the lungs and disposal of HCO3- as a consequence of ureagenesis in the liver.

Acid-Base Equilibrium↗
PubMed1985

Modulation of the activity of rat liver acetylglutamate synthase by pH and arginine concentration.

Acetylglutamate is known to modulate the activity of carbamyl phosphate synthetase, and thus probably to participate in regulation of the urea cycle. Therefore factors that regulate the activity of acetylglutamate synthase are relevant to control of urea synthesis and of systemic pH. An increase in the concentration of arginine increases both Vmax and S0.5 for glutamate of acetylglutamate synthase from rat liver. An increase in pH causes S0.5 for glutamate to decrease and does not affect Vmax. As a consequence of these effects, a rapid rate of synthesis of acetylglutamate requires a concentration of arginine of about 25 microM or higher and either relatively high glutamate concentrations or relatively high pH.

Acetyl Coenzyme A↗
PubMed1985

Interactions between citrate and nucleoside triphosphates in binding to phosphofructokinase.

Interrelationships between the binding by rabbit muscle phosphofructokinase of citrate, ATP, GTP, and adenyl-5'-yl imidodiphosphate (AMP-PNP) were investigated. To allow measurements at 25 degrees C, pyruvate kinase and phosphoenolpyruvate were included in the dialysis media to rephosphorylate ADP formed by the weak ATPase action of phosphofructokinase. Binding of citrate was enhanced by GTP nearly as much as by ATP, although GTP does not inhibit the catalytic activity of the enzyme. The results are consistent with the interpretation that binding of GTP, and, by analogy, ATP, at the catalytic site enhances the binding of citrate. AMP-PNP also enhanced citrate binding. Both ATP and GTP appear to bind at three sites per enzyme subunit, with the apparent third site binding relatively weakly. The estimated dissociation constants for the first two sites, about 33 microM for both for ATP compared with 3 and 280 microM for GTP, are consistent with kinetic results that imply lack of effective competition by GTP for the inhibitory site. When a compound binds at two or more sites on a macromolecule, the position and shape of the binding curve are sensitive to the geometric mean of the binding constants but quite insensitive to the magnitudes of the individual constants; thus, binding affinities cannot be estimated with confidence in such cases.

Adenosine Triphosphate↗