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Biomedical subjects

G K Radda

Publications and source records attributed to G K Radda.

At least 325 records · Page 18Linked to original sources
PubMed1982

The effects of insulin on myocardial metabolism and acidosis in normoxia and ischaemia. A 31P-NMR study.

1. The effect of insulin on the perfused rat heart during normoxia and total ischaemia was studied by 31P-NMR. 2. During normoxic perfusion, insulin increased the phosphocreatine to ATP ratio at the expense of Pi, when glucose was the substrate. No change was observed when acetate was used as the sole substrate. the intracellular pH (as measured from the position of the 2-deoxyglucose 6-phosphate resonance peak) was unaffected by insulin treatment. 3. Infusion of insulin prior to ischaemia caused an increase in the rate and extent of acidosis during the period of no flow while the rate of ATP depletion was decreased. 4. Freeze-clamped studies showed an increase in glycogen levels upon insulin treatment of the glucose perfused rat heart. During ischaemia, a decrease in glycogen content concomitant with an increase in lactate was observed. The accessibility of glycogen to phosphorylase during ischaemia is increased as a result of insulin treatment. The control of glycolysis during ischaemia is discussed with respect to the content and structure of glycogen in heart tissue.

Acidosis↗
PubMed1982

Oxygenation dependence of the transverse relaxation time of water protons in whole blood at high field.

At high and medium magnetic field, the transverse NMR relaxation rate (T-1(2)) of water proteins in blood is determined predominantly by the oxygenation state of haemoglobin. T-1(2) depends quadratically on the field strength and on the proportion of haemoglobin that is deoxygenated. Deoxygenation increases the volume magnetic susceptibility within the erythrocytes and thus creates local field gradients around these cells. From volume susceptibility measurements and the dependence of T-1(2) on the pulse rate in the Carr-Purcell-Meiboom-Gill experiment, we show that the increase in T-1(2) with increasing blood deoxygenation arises from diffusion of water through these field gradients.

Animals↗
PubMed1982

Energy metabolism and cellular pH in normal and pathological conditions. A new look through 31phosphorus nuclear magnetic resonance.

Cellular energetics can be studied non-invasively using 31P nuclear magnetic resonance (NMR). Besides the concentration of the major phosphorus-containing metabolites, enzyme-catalysed fluxes and intracellular pH can be obtained from the NMR measurement. Metabolic acidosis in the ischaemic myocardium has been studied in the perfused rat heart. Comparison of the pH, measured from the position of the inorganic phosphate (Pi) resonance, with the pH obtained from the resonance of 2-deoxyglucose 6-phosphate (accumulated after perfusion with 2-deoxyglucose) demonstrates that cytoplasmic pH values are being obtained. The pH changes during ischaemia are quantitatively related to the glycogen leads to lactate conversion. Preperfusion with insulin enhanced the lactate production but resulted in the maintenance of ATP for a longer period during ischaemia, even though acidosis was enhanced. In acute renal acidosis, the decrease in intracellular pH is smaller than might have been expected, as demonstrated in the perfused rat kidney. The importance of acidosis in relation to renal preservation was examined in human kidneys prior to transplantation. 31P NMR studies on human forearm muscle have been done. The pH changes measured during aerobic and anaerobic exercise demonstrate the importance of glycogenolysis in providing energy for ATP production. In a patient with suspected McArdle's syndrome, 31P NMR was used to detect the lack of glycogenolysis since during both aerobic and anaerobic exercise a pH rise was observed together with the rapid breakdown of phosphocreatine.

Acidosis↗
PubMed1982

Preliminary observations on the metabolic responses to exercise in humans, using 31-phosphorus nuclear magnetic resonance.

The new non-invasive method of 31P nuclear magnetic resonance (NMR) has been applied for the first time to human muscle. The metabolic response to exercise--a fall in intracellular pH, a fall in phosphocreatine content, and an increase in inorganic phosphate (Pi) content--occurs without any change in ATP content of the exercised muscle. Abnormal spectra occur in two myopathies examined to date: in McArdle's syndrome, pH rises during exercise, in contrast to the normal fall; and, in an unusual mitochondrial myopathy, Pi content is high, relative to phosphocreatine content, and in keeping with an excessive oxygen consumption in this patient. Intracellular pH reflected, in addition, the systemic pH of the subject; the anticipated abnormalities in muscle pH have been observed in association with lactic acidosis, renal failure and hyperventilation.

Acidosis, Respiratory↗
PubMed1981

A 31P-NMR study of the effects of reflow on the ischaemic rat heart.

(1) The recovery of perfused rat hearts experiencing various lengths of total global ischaemia was studied using 31P-NMR. Mechanical function was monitored by measuring left ventricular pressure. (2) Hearts exposed to a maximum of 14 min total global ischaemia regained stable contractile function on reperfusion. The concentration of phosphocreatine in these hearts rapidly exceeded its pre-ischaemic value while that of ATP rose very slowly. Pi fell on reflow to approximately its original level. These observations are interpreted as being the result of a rapid turnover of ATP stimulating phosphocreatine production by the mitochondrial isozyme of creatine kinase (ATP: creatine N-phosphotransferase, EC 2.7.3.2). (3) The recovery of intracellular pH on reperfusion does not depend upon the duration of ischaemia, nor on the pH or the percentage of ATP depletion at the end of the ischaemic period. This indicates that pH recovery is a flow-dependent phenomenon. (4) In non-recovering hearts, multiple Pi resonances are observed which arise from areas of differing myocardial pH. Phosphocreatine levels did not rise above 50% of their pre-ischaemic values. ATP levels remained depressed. This suggests that localized tissue necrosis only characterizes the failing situation.

Adenosine Triphosphate↗
PubMed1981

The role of phospholipids in the modulation of enzyme activities in the chromaffin granule membrane.

(1) 93% of protein of chromaffin granule membranes can be solubilized by 1.3% (w/v) sodium cholate. The solubilized material can be substantially delipidated by ammonium sulphate precipitation. After three such cycles less than 2% of the endogenous phospholipids remain. (2) The chromaffin granule membrane Mg2+-ATPase depends on the presence of phospholipids for retention of its full activity. Soybean and extracted chromaffin granule phospholipids fully reactivate the delipidated enzyme provided only one delipidation step is used. (3) Successive ammonium sulphate precipitation steps result in a delipidated, and deactivated ATPase preparation which can be only partially reactivated on re-addition of phospholipids. (4) The phospholipid specificity for reactivation of the Mg2+-ATPase is broad. Although acidic phospholipids allow higher activities than neutral phospholipids, the main requirement appears to be the hydrophobic environment provided by the phospholipid hydrocarbon chains. (5) Correlations between changes in slope in the Arrhenius plot of the Mg2+-ATPase, and phase transitions in the phospholipid used for reactivation suggest that the 'fluidity' of the hydrocarbon chains modulates the activity of the enzyme.

Adenosine Triphosphatases↗
PubMed1981

The effect of fatty acids on the surface potential of phospholipid vesicles measured by condensed phase radioluminescence.

(1) The carboxyl group of fatty acids has a very low pK value, which is shifted into the physiological pH range when they are incorporated in a phospholipid membrane. As a result of a pH increase the surface charge and surface potential of the membrane increase. (2) The titration of the carboxyl group was observed with condensed phase radioluminescence. This technique uses the electron emitted by the tritiated membrane probe (oleic acid or cholesterol) to excite a fluorophore also incorporated in the bilayer. (3) The phase transition of dimyristoyl phosphatidylcholine vesicles labelled within 12-(9-anthroyloxy)stearic acid was measured by condensed phase radioluminescence at different pH values. (4) We related the condensed phase radioluminescence signal to the asymmetrical distribution of the fluorophore between the inner and outer layer of the lipid membrane which is induced by the repulsion of the negatively charged fatty acids. (5) We showed that the condensed phase radioluminescence signal is proportional to the protonation of the carboxyl group. On this basis, the broadening of the titration curve can be explained as an effect of the self-induced membrane potential calculated using the Gouy-Chapman theory. (6) Ca2+ drastically reduces the flip-flop rate of fatty acids across the membrane and also caused a decrease in the asymmetric distribution. (7) We concluded that a fatty acid can act as a membrane surface buffer. The pK value of 12-(9-anthroyloxy)stearic acid in a dimyristoyl phosphatidylcholine membrane is 7.0 +/- 0.3. (8) We discuss the results with respect to aggregation, fusion and clustering.

Cholesterol↗
PubMed1981

Activity of phosphorylase in total global ischaemia in the rat heart. A phosphorus-31 nuclear-magnetic-resonance study.

1. The uptake and subsequent phosphorylation of deoxyglucose into perfused rat hearts was monitored by 31P n.m.r. 2. The accumulated deoxyglucose 6-phosphate provided (a) an independent method for measuring cytosolic pH in the normoxic and ischaemic heart tissue and (b) a way of studying the activity of phosphorylase during ischaemia. 3. The cytosolic pH measured from the 31P n.m.r. resonance position of deoxyglucose 6-phosphate is in good agreement under all conditions studied with that obtained previously from the Pi resonances. This eliminates any possible doubts about the use of Pi for measuring intracellular pH. 4. Deoxyglucose 6-phosphate in vitro inhibits phosphorylase b but not phosphorylase a. Its inhibitory effect on glycogenolysis during ischaemia is monitored by measuring tissue acidosis by n.m.r. In the initial stages of ischaemia phosphorylase activity is not inhibited, whereas after about 5 min approx. 50% of the activity is inhibited. These observations are interpreted in terms of the relative contributions of phosphorylase a and the AMP-dependent phosphorylase b activities during ischaemia.

Animals↗
PubMed1981

The activity of creatine kinase in frog skeletal muscle studied by saturation-transfer nuclear magnetic resonance.

1. The activity of creatine kinase in intact anaerobic frog muscle at 4 degrees C at rest and during contraction was investigated by using saturation-transfer 31P n.m.r. 2. At rest, the measured forward (phosphocreatine to ATP) reaction flux was 1.7 X 10(-3) M . s-1 and the backward flux was 1.2 X 10(-3) M . s-1. The large magnitude of both fluxes shows that creatine kinase is active in resting muscle, so the observed constancy of [phosphocreatine] demonstrates that the enzyme and its substrates are at equilibrium. 3. The apparent discrepancy between the fluxes must arise largely from an underestimation of the backward flux resulting from interaction of ATP with other systems, e.g. via adenylate kinase. For purposes of further calculation we have therefore adopted 1.6 X 10(-3) M . s-1 as an estimate of both fluxes. 4. During contraction, when the creatine kinase reaction is no longer at equilibrium, the net rate of phosphocreatine breakdown, estimated directly from the change in area of the inorganic phosphate peak, was 0.75 X 10(-3) M . s-1. Saturation transfer indicates that the forward reaction flux remains at approx. 1.6 X 10(-3) M . s-1 and the backward flux decreases to about 0.85 X 10(-3) M . s-1. 5. The activity of creatine kinase during contraction is large enough to account for the well-established observation that, during contraction, the concentration of ATP falls by less than 2-3%. The reaction catalysed by creatine kinase is driven forward during contraction by the large relative increase in the concentration of free ADP, which is more than doubled. 6. The observation that the forward flux does not increase during contraction and that the backward flux decreases can most simply be explained on the basis of competition of reactants for a limited amount of enzyme.

Adenosine Triphosphate↗
PubMed1981

Phosphorus nuclear magnetic resonance study of the rat kidney in vivo.

We show that phosphorus nuclear magnetic resonance (31P-NMR) can be used to study the metabolic state of kidneys in live, anesthetized rats without the need for surgery. To localize signals from the kidney, we used a radiofrequency surface coil in conjunction with the magnetic field profiling technique that is used for "topical magnetic resonance," (TMR). Signals are observed from phosphorus-containing metabolites including ATP and inorganic phosphate, and under certain conditions intracellular pH can be estimated. We suggest that the ratio of free ATP to free ADP is higher than the estimates of 1.5 to 2.0 obtained from freeze-clamping studies. Because of the important roles that ATP, ADP, inorganic phosphate, and pH are believed to play in cellular metabolism, it seems that 31P-NMR could be a powerful clinical tool.

Adenosine Triphosphate↗
PubMed1981

The role of intrarenal pH in regulation of ammoniagenesis: [31P]NMR studies of the isolated perfused rat kidney.

1. [31P]NMR spectra were obtained from a functioning isolated perfused rat kidney with the aim of determining intrarenal pH in acute acidosis. 2. Signals from intracellular inorganic phosphate could be observed in the absence of phosphate in the perfusion medium. Under these conditions renal ATP and inorganic phosphate content fell by 30% but total adenine nucleotide and phosphorylation potential ATP/ADP x Pi were unchanged compared with kidneys perfused with phosphate-containing medium. In addition, G.F.R., Na+ reabsorption and ammonia formation from glutamine remained normal. Ammonia production increased 93%, urine pH fell to 5.8 +/- 0.1 and kidney 2-oxoglutarate content fell by 80% upon acidification of the perfusion medium from pH 7.4 to pH 6.9, findings identical with those obtained in controls (Ross & Tannen, 1979). 3. [31P]NMR spectra of the isolated perfused rat kidney showed a pattern of adenine nucleotides and a small concentration of phosphocreatine, Intra-renal pH was measured from the resonance position of intracellular inorganic phosphate and in perfusions with pH 7.4 buffer was 7.19 +/- 0.10 (n = 11). 4. Acidification of the perfusion medium to pH 7.0 resulted in 0.3 pH unit fall in intrarenal pH. This fall in total intrarenal pH is insufficient to explain the fall in 2-oxoglutarate concentration observed if the glutamate-dehydrogenase-equilibrium model is invoked. 5. The line-width of the NMR signal is compatible either with heterogeneity of intra-renal pH or the existence of a pH gradient between cytosol and mitochondria, or both.

Acidosis↗
PubMed1981

NMR studies of tissue metabolism.

During the past six or seven years, high resolution nuclear magnetic resonance (NMR) has been established as a powerful tool for the study of cellular metabolism in isolated cells and organs. The non-invasive nature of the measurement offers many advantages over conventional analytical techniques, but limitations on sensitivity restrict the method to the observation of molecules present in the cell at relatively high concentrations (in the order of 0.5-1 mM). Inevitably, many of the initial investigations tended to confirm existing knowledge, but this was important in establishing the validity of the NMR method. In the last few years, NMR has contributed new and interesting information on several aspects of metabolism and metabolic regulation. The majority of reports deal with 31P, 13C, and, to a lesser extent, 1H NMR. The particular areas of interest include measurements of metabolic fluxes in the steady state and transport processes, observations of cellular pH and H+ ion distribution, and compartmentation and spatially resolved NMR of whole animals. The ability to directly observe the intracellular milieu is perhaps the major single advantage of NMR. A very large number of different systems can be studied, but it is important to keep in mind that NMR is simply an additional method for studying metabolism and must always be combined with carefully designed biochemical experiments.

Adenine Nucleotides↗