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G J Barritt

Publications and source records attributed to G J Barritt.

At least 91 records · Page 5Linked to original sources
PubMed1981

Evidence for two compartments of exchangeable calcium in isolated rat liver mitochondria obtained using a 45Ca exchange technique in the presence of magnesium, phosphate, and ATPase at 37 degrees C.

The distribution of calcium between isolated rat liver mitochondria and the extramitochondrial medium at 37 degrees C and in the presence of 2 mM inorganic phosphate, 3 mM ATP, 0.05 or 1.1 mM free magnesium and a calcium buffer, nitrilotriacetic acid, was investigated using a 45Ca exchange technique. The amounts of 40Ca in the mitochondria and medium were allowed to reach equilibrium before initiation of the measurement of 45Ca exchange. At 0.05 mM free magnesium and initial extramitochondrial free calcium concentrations of between 0.15 and 0.5 microM, the mitochondria accumulated calcium until the extramitochondrial free calcium concentration was reduced to 0.15 microM. Control experiments showed that the mitochondria were stable under the incubation conditions employed. The 45Ca exchange data were found to be consistent with a system in which two compartments of exchangeable calcium are associated with the mitochondria. Changes in the concentration of inorganic phosphate did not significantly affect the 45Ca exchange curves, whereas an increase in the concentration of free magnesium inhibited exchange. The maximum rate of calcium outflow from the mitochondria was estimated to be 1.7 nmol/min per mg of protein, and the value of K0.5 for intramitochondrial exchangeable calcium to be about 1.6 nmol per mg of protein. Ruthenium Red decreased the fractional transfer rate for calcium inflow to the mitochondria while nupercaine affected principally the fractional transfer rates for the transfer of calcium between the two mitochondrial compartments. The use of the incubation conditions and 45Ca exchange technique described in this report for studies of the effects of agents which may alter mitochondrial calcium uptake or release (e.g., the pre-treatment of cells with hormones) is briefly discussed.

Adenosine Triphosphate↗
PubMed1981

A kinetic analysis of the effects of adrenaline on calcium distribution in isolated rat liver parenchymal cells.

1. The effects of adrenaline on Ca distribution in isolated rat liver parenchymal cells were studied using a (45)Ca exchange technique under steady-state conditions with respect to the net movement of Ca. (45)Ca was initially introduced into the extracellular medium. The amount of cellular (45)Ca was determined after separation of the cells from the medium by centrifugation through a solution which contained LaCl(3) (to displace (45)Ca bound to sites on the outside of the cell membrane) and silicon oil. At 1.3 and 2.4 mm-extracellular Ca, a stimulation of the initial rate of (45)Ca exchange was observed in the presence of 10(-7)m-adrenaline (or 10(-6)m-phenylephrine) with a 7% decrease, and no change, respectively, in the plateau of the exchange curve. The same degree of stimulation was observed when (45)Ca was added at 1, 15, 30 or 45 min after the adrenaline.2. No stimulation of the initial rate of exchange was observed at 0.1 mm-extracellular Ca, or at 2.4 mm-extracellular Ca in the presence of antimycin A and oligomycin. At 0.1 mm-Ca, a 60% decrease in the plateau of the exchange curve was observed in the presence of adrenaline. The concentration of adrenaline (10(-7)m) which caused half-maximal stimulation of the initial rate of (45)Ca exchange at 1.3 mm-Ca was similar to that (2 x 10(-7)m) which caused half-maximal decrease in the plateau at 0.1 mm-Ca.3. The addition of adrenaline to cells equilibrated with (45)Ca at either 2.4 or 1.3 mm-Ca caused a transient loss of (45)Ca followed by a return to a new steady state after 1 or 10 min, respectively. A loss of (45)Ca was also observed at 0.1 mm-Ca, but the (45)Ca content of the cells remained maximally depressed for at least 30 min.4. A non-linear least-squares iterative curve-fitting technique was used to demonstrate that (a) an equation which includes two exponential terms and (b) a parallel or series arrangement of three compartments of exchangeable Ca (the medium and two compartments associated with the cell) are consistent with each set of data obtained at 1.3 or 2.4 mm-Ca in the presence or absence of adrenaline (or phenylephrine). At 1.3 mm-Ca, the quantities of exchangeable Ca in the two kinetically defined cellular compartments were 0.04-0.07 and 0.34-0.37 nmol per mg wet weight with rate constants for Ca outflow of 1.2-1.5 and 0.06-0.08 min(-1), respectively.5. Analysis of the changes induced by adrenaline or phenylephrine showed that at 1.3 and 2.4 mm-extracellular Ca these agents caused a 75-150% increase in the quantity of exchangeable Ca in the small kinetically defined compartment and a 20% decrease in the quantity of exchangeable Ca in the large kinetically defined compartment. These changes were mediated by an 80-160% increase in the rate constant for the inflow of Ca from the medium to the small kinetically defined compartment, and either a 20-60% decrease in the rate constant for inflow to, or a 20% increase in the rate constant for outflow from, the large compartment.6. Replacement of the LaCl(3) in the solution used to separate the cells from the incubation medium with either 5 mm-EGTA or 5 mm-CaCl(2) did not alter the kinetics of (45)Ca exchange or the stimulation by adrenaline. This, together with the observation that at 1.3 mm-extracellular Ca, adrenaline increases the initial rate of exchange in the absence, but not in the presence, of antimycin A plus oligomycin, indicates that both cellular compartments of exchangeable Ca are intracellular.7. The addition of antimycin A plus oligomycin to cells equilibrated with (45)Ca at 2.4 mm-extracellular Ca in the presence or absence of adrenaline displaced 0.09 and 0.14 nmol (45)Ca. mg(-1), respectively.8. Subcellular fractionation of cells equilibrated with (45)Ca at 0.1 mm-extracellular Ca revealed that the mitochondria and microsomes contained significant amounts of (45)Ca. The amounts of (45)Ca in these fractions decreased by 50 and 40%, respectively, in the presence of adrenaline.9. In (45)Ca exchange experiments conducted with isolated mitochondria at 37 degrees C at 1.5 x 10(-7)m and 0.9 x 10(-7)m free Ca in the presence of 2 mm-Mg(2+), one kinetically defined compartment of exchangeable mitochondrial Ca was detected. The rate constants for Ca outflow were found to be 0.15+/-0.03 and 0.12+/-0.04 min(-1), respectively, in reasonable agreement with the value obtained for the rate constant for the outflow of Ca from the large kinetically defined compartment of exchangeable Ca observed in cells.10. It is concluded that adrenaline has two effects on Ca movement in the liver cell. These are to cause a loss of Ca from an intracellular compartment, which includes the mitochondria and microsomes, and to increase the transport of Ca from the extracellular medium to an intracellular site. This results in an increase in the amount of Ca in a small intracellular compartment which may represent cytoplasmic Ca, or Ca bound to sites on the inside of the plasma membrane.

Animals↗
PubMed1979

Interaction between glucocorticoids and glucagon in the hormonal modification of calcium retention by isolated rat liver mitochondria.

1. The administration of dexamethasone to intact fed rats by intraperitoneal injection for 3h was associated with a 6-fold increase in the time for which mitochondria subsequently isolated from the liver retain a given load of exogenous Ca2+. This effect was blocked by the co-administration of cycloheximide with dexamethasone, and partially blocked by the co-administration of puromycin. Daily administration of dexamethasone for periods of 4--7 days resulted in liver mitochondria that exhibited a decreased ability to retain exogenous Ca2+. 2. When glucagon was administered to fed adrenalectomized rats, the increase in mitochondrial Ca2+-retention time that results from the action of this hormone was reduced by 50% when compared with its effect on intact animals. The administration of dexamethasone to adrenalectomized rats partially restored the full effect of glucagon. 3. Dexamethasone did not enhance the effect of glucagon on mitochondrial Ca2+-retention time when administered to intact fed rats. 4. It is concluded that these data support the hypothesis that the hormone-induced modification of liver mitochondria, which results in an increase in the time for which exogenous Ca2+ is retained, involves a step in which new protein is synthesized.

Adrenalectomy↗
PubMed1978

Effects of glucagon and N6O2'-dibutyryladenosine 3':5'-cyclic monophosphate on calcium transport in isolated rat liver mitochondria.

1. The administration of glucagon to fed rats by intraperitoneal injection, or the perfusion of livers from fed rats with glucagon by the method of Mortimore [Mortimore (1963) Am.J. Physiol. 204, 699--704] was associated with increases of 15- and 5-fold respectively, in the time for which a given load of exogenous Ca2+ is retained by mitochondria subsequently isolated from the liver. This effect of glucagon was (a) also induced by N6O2'-dibutyryl cyclic AMP, (b) completely blocked by cycloheximide, (c) relatively slow in onset (15--60 min) and (d) associated with a stimulation of about 20% in the rates of ADP-stimulated oxygen utilization and Ca2+ transport measured in the presence of succinate. 2. Perfusion of livers with glucagon resulted in the isolation of mitochandria which showed a 50% increase, no significant change and a 40% increase in the concentrations of endogenous Ca, Mg and Pi respectively, when compared with mitochondria isolated from control perfused livers. 3. The administration of insulin or adrenaline to fed rats induced increases of 10- and 8-fold respectively, in the time for which Ca2+ is retained by isolated liver mitochondria. Perfusion of livers with insulin had no effect on mitochondrial Ca2+ retention time. 4. The perfusion of livers from starved rats with glucagon, or the administration of either glucagon or insulin to starved rats, increased by about 2.5- and 15-fold respectively, the time for which isolated mitochondria retain Ca2+. 5. Mechanisms which may be responsible for the observed alterations in Ca2+-retention time are discussed.

Animals↗
PubMed1978

Effects of hormones and N6O2'-dibutyryl-adenosine 3' :5'-cyclic monophosphate, administered in vivo, on phosphate transport and metabolism in isolated rat liver mitochondria.

1. The administration of glucagon or N6O2'-dibutyryl cyclic AMP to fed rats by intraperitoneal injection was associated with a 2-fold increase in the amounts of endogenous Pi and ATP, and an increase in the rate and extent of transport of exogenous Pi (measured in either the presence or the absence of Ca2+) in mitochondria subsequently isolated from the liver. No change was observed in either the maximum rate of transport of exogenous Pi or in the rate of 32Pi exchange. 2. The changes induced by glucagon and dibutyryl cyclic AMP were markedly decreased by the co-administration of cycloheximide. 3. The administration of insulin to rats resulted in an increase of about 1.3-fold in the concentration of endogenous mitochondrial Pi 4. The amounts of endogenous Pi in mitochondrial isolated from the livers of starved rats were 3 times those in mitochondria isolated from fed animals. 5. It is concluded that the liver mitochondrial phosphatetransport system may be an important site of hormone action. 6. In the course of these experiments, it was shown that Ca2+ markedly stimulates mitochondrial phosphate transports.

Animals↗
PubMed1975

The effect of butacaine on adenine nucleotide binding and translocation in rat liver mitochondria.

The effect of the local anaesthetic, butacaine, on adenine nucleotide binding and translocation in rat liver mitochondria partially depleted of their adenine nucleotide content was investigated. The range of butacaine concentrations that inhibit adenine nucleotide translocation and the extent of the inhibition are similar to the values obtained for native mitochondria. Butacaine does not alter either the total number of atractyloside-sensitive binding sites of depleted mitochondria, or the affinity of these sites for ADP or ATP under conditions where a partial inhibition of the rate of adenine nucleotide translocation is observed. The data are consistent with an effect of butacaine on the process by which adenine nucleotides are transported across the mitochondrial inner membrane rather than on the binding of adenine nucleotides to sites on the adenine nucleotide carrier. The results are briefly discussed in relation to the use of local anaesthetics in investigations of the mechanism of adenine nucleotide translocation.

4-Aminobenzoic Acid↗
PubMed1975

Stimulation of hepatic mitochondrial calcium transport by elevated plasma insulin concentrations.

The effect of insulin (injected intraperitoneally) on the transport of Ca2+ by hepatic mitochondria from rats was investigated. 2. Elevated concentrations of plasma insulin within the physiological range (10-100muunits/ml) stimulate the initial rate of Ca2+ transport into mitochondria at 4 degrees C by about 75% and prolong by approx. tenfold the time for which the mitochondria retain the accumulated Ca2+. 3. The prolonged retention of Ca2+ is observed under the conditions where hypoglycaemia is significantly decreased by the simultaneous injection of glucose and insulin. 4. A good correlation is observed between the effects on Ca2+ transport and the decrease in blood glucose concentration when the amount of insulin injected was varied. 5. The effects of insulin on mitochondrial Ca2+ transport are apparent at about 30 min after the injection, and are inhibited by cycloheximide. 6. There is little change in mitochondrial energy transduction after the administration of insulin. 7. The results are briefly discussed in relation to the mechanisms of Ca2+ transport across the inner mitochondrial membrane and the role of mitochondria in modifying intracellular Ca2+ concentrations with reference to the mechanism(s) by which insulin affects cellular metabolism.

Animals↗