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

Abstract

SUMMARY1. The effects of adrenaline on Ca distribution in isolated rat liver parenchymal cells were studied using a 45Ca exchange technique under steady-state conditions with respect to the net movement ofCa. 45Ca was initially introduced into the extracellular medium. The amount of cellular 45Ca was determined after separation ofthe cells from the medium by centrifugation through a solution which contained LaCl3 (to displace 45Ca 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 45Ca 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 45Ca 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 mMextracellular 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 45Ca 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 45Ca at either 2-4 or 1-3 mM-Ca caused a transient loss of 45Ca followed by a return to a new steady state after 1 or 10 min, respectively. A loss of 45Ca was also observed at 0-1 mM-Ca, but the 45Ca 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 ofadrenaline (or phenylephrine). G. J. BARRITT, J. C. PARKER AND J. C. WADSWORTH At 1P3 mM-Ca, the quantities of exchangeable Ca in the two kinetically defined cellular compartments were 0-04-0-07 and 034-4037 nmol per mg wet weight with rate constants for Ca outflow of 1P2-1P5 and 0-06-0-08 min-', 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 ofexchangeable 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 LaCl3 in the solution...

Calcium: its modulation in liver by cross-talk between the actions of glucagon and calcium-mobilizing agonists

“…The main methods employed to measure rates of Ca2+ inflow are the use of 45Ca under steady-state conditions (see Barritt et al, 1981), (indirect) measurement of increases in intracellular Ca2+ using fluorescent indicators following the addition of Ca2+ to a medium containing negligible Ca2 , measurement of the rates of quenching of fluorescence of intracellular quin2 following Mn2+ addition (see, e.g., Crofts and Barritt, 1990), and measurement of the initial rate of activation of glycogen phosphorylase following the addition of Ca2+ to a medium containing negligible Ca2+. A further complication to studies of Ca2+ inflow is that there is mounting evidence that more than one system facilitates Ca2+ entry in the plasma membrane of liver cells (Barritt et al, 1981;Hughes et al, 1986;Altin and Bygrave, 1987a;Barritt and Hughes, 1991;Llopis et al, 1992). The potentially useful technique of patch-clamping appears to be technically difficult in liver plasma membranes (see, e.g., Sawanobori et al, 1989;Barritt and Hughes, 1991).…”

Calcium and long‐term transmission damage following anoxia in dentate gyrus and CA1 regions of the rat hippocampal slice.

“…This equation has been applied to 45Ca21 uptake into a variety of tissues including smooth muscle (Scheid & Fay, 1984), liver cells (Barritt, Parker & Wadsworth, 1981) and cultured kidney cells (Borle, 1970). In the derivation, 45Ca2+ uptake is assumed to take place from an effectively infinite pool into two exchangeable cellular pools whose sizes are Al and A2 and whose first-order efflux rate constants are K1 and K2 (Borle, 1975 (Borle, 1975).…”

Exogenous phospholipase enzymes mimic effects of phenylephrine on Ca2+ transport in hepatocytes

“…A large body of evidence indicates that this is due to the release of Ca2+ from the mitochondria and endoplasmic reticulum (Chen et al, 1978;Babcock et al, 1979;Blackmore et al, 1979;Murphy et al, 1980;Barritt et al, 1981b;Berthon et al, 1981) and enhancement of Ca2+ inflow across the plasma membrane (Keppens et al, 1977;Assimacopoulos-Jeannet et al, 1977;Foden & Randle. 1978;Barritt et al, 1981b).…”

“…A large body of evidence indicates that this is due to the release of Ca2+ from the mitochondria and endoplasmic reticulum (Chen et al, 1978;Babcock et al, 1979;Blackmore et al, 1979;Murphy et al, 1980;Barritt et al, 1981b;Berthon et al, 1981) and enhancement of Ca2+ inflow across the plasma membrane (Keppens et al, 1977;Assimacopoulos-Jeannet et al, 1977;Foden & Randle. 1978;Barritt et al, 1981b). It has been postulated that metabolites formed during hydrolysis of plasma membrane phosphatidylinositol catalyse the inflow of Ca2+ across the plasma membrane (Michell, 1975: Billah & Michell, 1979Barritt et al, 1981a) (but see Hawthorne, 1982) and its release from intracellular stores (Billah & Michell, 1979: Barritt, 1981: Whiting & Barritt, 1982.…”