Calcium: its modulation in liver by cross-talk between the actions of glucagon and calcium-mobilizing agonists.
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Animals↗
Biomedical subjects
F L Bygrave
Publications and source records attributed to F L Bygrave.
Evidence that stimulation of plasma-membrane Ca2+ inflow is an early action of glucagon and dibutyryl cyclic AMP in rat hepatocytes.
The ability of glucagon (1 nM) and of dibutyryl cyclic AMP (50 microM) to increase cytosolic free Ca2+ concentration ([Ca2+]i) in Fura-loaded rat hepatocytes was examined in a system wherein Ca2+ inflow was induced by the re-admission of excess Ca2+ to a nominally Ca(2+)-free medium. An increase in [Ca2+]i did not occur in the absence of either agonist, but did so after co-addition of either agonist with Ca2+. Increasing the time between addition of dibutyryl cyclic AMP (or of glucagon) and Ca2+ led to increases in [Ca2+]i; half-maximal and maximal increases were observed at 0 s (i.e. at co-addition) and 5-7 s respectively. Dibutyryl cyclic AMP and Ca2+ each exhibited a concentration-dependence when their respective concentrations were changed for a fixed time interval between additions. Half-maximal and maximal effects were obtained with 30 microM and 50 microM dibutyryl cyclic AMP and with 0.5 mM and approx. 1 mM Ca2+ respectively. The data demonstrate an early action of glucagon and dibutyryl cyclic AMP on [Ca2+]i. It is argued that the agonist-induced rise in [Ca2+]i results from an increase in plasma-membrane Ca2+ inflow, an effect that appears to occur much earlier than that on mobilization of internal stores of Ca2+.
Animals↗
Different 1,4-dihydropyridines exhibit discriminating effects on passive calcium uptake in rat liver plasma membrane vesicles.
The effects of a number of calcium channel effectors on Ca2+ uptake by rat liver plasma membrane vesicles was examined. Nifedipine, verapamil and diltiazem had to be present at 1 mM in order to produce > 50% inhibition of Ca2+ uptake. The two structurally similar 1,4-dihydropyridines, nicardipine and nisoldipine exhibited opposite effects; nicardipine inhibited while nisoldipine stimulated Ca2+ uptake. The results show that low concentrations (microM) of calcium channel blockers of excitable cells have little effect on Ca2+ uptake by liver plasma membrane vesicles consistent with earlier findings of others that voltage-gated calcium channels are absent in hepatocytes. However, the opposite effects of higher concentrations (ca. 1 mM) of nicardipine and nisoldipine on Ca2+ uptake suggest a discriminatory action that might be useful in studying further the mechanism of passive Ca2+ uptake by these membrane vesicles.
Animals↗
Restoration of mitochondrial energy-linked reactions following dexamethasone treatment of rats infected with the liver fluke Fasciola hepatica.
Mitochondria isolated from male Wistar rats experimentally infected with the common liver fluke, Fasciola hepatica, exhibit loss of respiratory control from 2 weeks post-infection (Rule, et al. (1989) Biochem. J. 260, 517-523). We now report that subcutaneous injections of the anti-inflammatory drug, dexamethasone, during the final week of infection prevented the mitochondrial uncoupling and restored respiratory control almost to the levels of uninfected controls. Further investigations have shown that mitochondria from infected rat livers are unable to synthesize ATP and that abnormal respiration is also evident in hepatocytes isolated from infected rats. These abnormalities were absent when infected rats were treated with dexamethasone. In addition, liver mitochondrial function in infected, congenitally athymic, nude rats (CBH/R nu/nu) was not significantly different from that in uninfected nude or Wistar controls. These results provide evidence that the mitochondrial dysfunction in fascioliasis is host-mediated and that T lymphocytes in particular may be involved.
Adenosine Triphosphate↗
Acute effects of cholestatic and choleretic bile salts on vasopressin- and glucagon-induced hepato-biliary calcium fluxes in the perfused rat liver.
The effects were investigated of the choleretic bile salt glycoursodeoxycholate (G-UDCA) and of the cholestatic bile salt taurochenodeoxycholate (T-CDCA) on changes in perfusate Ca2+, glucose and oxygen and in bile calcium and bile flow induced by the administration of (a) vasopressin, (b) glucagon and (c) glucagon plus vasopressin together to the perfused rat liver [Hamada, Karjalainen, Setchell, Millard & Bygrave (1992) Biochem. J. 281, 387-392]. G-UDCA itself increased the secretion of calcium in the bile several-fold, but its principal effect was to augment each of the above-mentioned metabolic events except glucose and oxygen output; particularly noteworthy was its ability to augment the 'transients' in bile calcium and bile flow seen immediately after the administration of vasopressin with or without glucagon. T-CDCA, by contrast, produced opposite effects and attenuated all of the parameters measured, and in particular the transients in bile calcium and bile flow. The data provide evidence of a strong correlation between calcium fluxes occurring on both the sinusoidal and the bile-canalicular membranes and that all are modifiable by glucagon, Ca(2+)-mobilizing hormones and bile salts.
Animals↗
Concomitant stimulation by vasopressin of biliary and perfusate calcium fluxes in the perfused rat liver.
Changes in perfusate Ca2+ (measured with a Ca(2+)-selective electrode) and changes in bile calcium (measured by atomic absorption spectroscopy) were continuously and simultaneously monitored after infusion of (a) vasopressin, (b) glucagon and (c) both vasopressin and glucagon together to the perfused rat liver. Also monitored were perfusate glucose and oxygen concentrations and bile flow. Vasopressin induces a sharp, transient, pulse of increased bile flow and increased bile calcium within 1 min of infusion, concomitant with rapid changes in perfusate Ca2+ fluxes, glucose output and oxygen uptake. This is immediately followed by a decrease in both bile flow and bile calcium for as long as the hormone is administered. Changes induced by glucagon are a relatively slow onset of perfusate Ca2+ efflux and oxygen uptake, but rapid glucose output, and a small but significant and transient decrease in bile flow and bile calcium which, despite the continued infusion of the hormone, spontaneously and rapidly returns to normality. However, the greatest responses are observed after co-administration of both hormones. Coincident with the augmented perfusate Ca2+ fluxes (influx) seen in earlier work, there occurs within 1 min of vasopressin infusion a sharp increase in bile secretion and bile calcium greater in magnitude than that produced by vasopressin alone. Immediately thereafter bile secretion and bile calcium decline below basal values and remain there for as long as the hormones are administered. Glucagon and vasopressin therefore each have opposing effects on bile flow and bile calcium. However, the action of vasopressin is enhanced by the prior administration of glucagon. The data thus reveal features about the actions of glucagon and Ca(2+)-mobilizing hormones on bile flow and bile calcium not previously recorded and provide a novel framework around which the whole issue of hepato-biliary Ca2+ homoeostasis can be assessed in normal and diseased liver.
Animals↗
Beneficial effect of dexamethasone on attenuated hormone-induced uptake of calcium and glycogenolysis by perfused liver of rats infected with Fasciola hepatica.
Infection of rats with the liver fluke, Fasciola hepatica, impaired the responses of the perfused liver to calcium uptake and glucose release induced by the synergistic action of glucagon and vasopressin. Treatment of infected rats with dexamethasone prevented the impairment of each of these two responses.
Animals↗
Aberrant energy-linked reactions in mitochondria isolated from the livers of sheep infected with the liver fluke Fasciola hepatica.
Respiration by mitochondria isolated from the livers of sheep following infection up to 15 weeks with F. hepatica was measured with the respiratory substrates pyruvate (plus malate) and succinate in the absence and presence of ADP; the rates were compared with those obtained by mitochondria isolated from livers of uninfected sheep. It was found that respiration supported by both substrates in mitochondria isolated from the left lobe but not the middle lobe of 4-week infected sheep exhibited abnormalities such that the acceptor control ratios were only marginally above one. Some, but not total, recovery was seen in the later stages of infection. The aberrant respiratory behaviour is similar to that observed with infected rats.
Animals↗
Phosphate and calcium uptake by mitochondria and by perfused rat liver induced by the synergistic action of glucagon and vasopressin.
Co-administration of glucagon and vasopressin to rat liver perfused with buffer containing 1.3 mM-Ca2+ induces a 4-fold increase in Pi in the subsequently isolated mitochondria (from approx. 9 to approx. 40 nmol/mg of mitochondrial protein). This increase is not attributable to PPi hydrolysis, and is not observed if the perfusate Ca2+ is lowered from 1.3 mM to 50 microM. The increase in mitochondrial Pi closely parallels that of mitochondrial Ca2+; when the increase in Pi and Ca2+ accumulation is maximal, the molar ratio is close to that in Ca3(PO4)2. Measurement of changes in the perfusate Pi revealed that, whereas administration of glucagon or vasopressin alone brought about a rapid decline in perfusate Pi, the largest decrease (reflecting net retention of Pi by the liver) was observed when the hormone was co-administered in the presence of 1.3 mM-Ca2+. The synergistic action of glucagon plus vasopressin was nullified by lowering the perfusate Ca2+ to 50 microM. The data provide evidence that, whereas glucagon may be able to alter Pi fluxes directly in intact liver, any alterations induced by vasopressin are indirect and result only from its action of mobilizing Ca2+.
Animals↗
Evidence for the involvement of carboxyl groups in passive calcium uptake by liver plasma membrane vesicles and in agonist-induced calcium uptake by hepatocytes.
The hydrophobic reagents DCCD and EEDQ, each of which reacts with protein carboxyl groups, were found to inhibit both passive Ca2+ uptake by plasma membrane vesicles isolated from rat liver and agonist-induced Ca2+ uptake by hepatocytes. The data raise the possibility that the Ca2+ inflow pathway(s) in liver has a specific requirement for a reactive carboxyl group or groups.
Animals↗
Aberrant energy-linked reactions in mitochondria isolated from the livers of rats infected with the liver fluke Fasciola hepatica.
The respiratory properties of mitochondria isolated from the livers of rats infected with the parasite Fasciola hepatica were examined. Oligomycin-sensitive ATPase activity was also examined during the acute stage (2-4 weeks post-infection). At 2,4 and 6 weeks post-infection, mitochondrial respiration in vitro (supported by site I and site II substrates) was completely uncoupled. Limited respiratory control had returned by 11 weeks post-infection, but complete recovery was not observed even at 21 weeks post-infection. At 4 weeks post-infection, uncoupled respiration (from all three energy-conserving sites) was also markedly attenuated (to the greatest extent with NADH-linked substrate). Except for pyruvate-supported respiration, this attenuation was not apparent at any other stage of the infection. The attenuation of pyruvate-supported respiration declined, but was still present, at 6 weeks post-infection. In addition to these perturbations in mitochondrial respiratory properties, mitochondrial ATPase activity at 4 weeks post-infection was insensitive to oligomycin, indicating a change in the structural integrity of the ATPase complex.
Adenosine Triphosphatases↗
Stimulation of hepatic inositol 1,4,5-trisphosphate kinase activity by Ca2+-dependent and -independent mechanisms.
A cytosolic fraction derived from rat hepatocytes was used to investigate the regulation of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] kinase, the enzyme which converts Ins(1,4,5)P3 to inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P4]. The activity was doubled by raising the free Ca2+ concentration of the assay medium from 0.1 microM to 1.0 microM. A 5 min preincubation of the hepatocytes with 100 microM-dibutyryl cyclic AMP (db.cAMP) plus 100 nM-tetradecanoylphorbol acetate (TPA) resulted in a 40% increase in Ins(1,4,5)P3 kinase activity when subsequently assayed at 0.1 microM-Ca2+. This effect was smaller at [Ca2+] greater than 0.5 microM, and absent at 1.0 microM-Ca2+. Similar results were obtained after preincubation with 100 microM-db.cAMP plus 300 nM-vasopressin (20% increase at 0.1 microM-Ca2+; no effect at 1.0 microM-Ca2+). Preincubation with vasopressin, db.cAMP or TPA alone did not alter Ins(1,4,5)P3 kinase activity. It is proposed that these results, together with recent evidence implicating Ins(1,3,4,5)P4 in the control of Ca2+ influx, could be relevant to earlier findings that hepatic Ca2+ uptake is synergistically stimulated by cyclic AMP analogues and vasopressin.
Animals↗
Exposure to depolarizing concentrations of K+ inhibits hormonally-induced calcium influx in rat liver.
The exposure of perfused rat livers to depolarizing concentrations of K+ (60 mM) by partial substitution of the NaCl in the medium with KCl induces glycogenolysis, respiratory changes and vasoconstriction. These responses were found to be inhibited 70-80% by 20 microM indomethacin and by 20 microM bromophenacyl bromide. This suggests that eicosanoids, namely prostaglandins, are involved in mediating these effects, and hence that the action of K+ involves primarily an effect on eicosanoid-producing cells (Kupffer and endothelial cells) within the liver. A 5 min pre-exposure of perfused livers to depolarizing concentrations of K+ (in the presence of indomethacin) was found to inhibit (by approx. 85%) the influx of Ca2+ induced by the co-administration of 10 nM glucagon and 10 nM vasopressin. A similar result was observed in isolated hepatocytes. The inhibition was probably not due to a decrease in the concentration of Na+ in the medium since the substitution of 80 mM NaCl with 80 mM choline chloride resulted in significantly less inhibition (30-40%). These results suggest that under these conditions the influx of Ca2+ in liver occurs through a pathway that is inhibited by high K+ concentration and/or a depolarization of the plasma membrane.
Acetophenones↗
Prostaglandin F2 alpha and the thromboxane A2 analogue ONO-11113 stimulate Ca2+ fluxes and other physiological responses in rat liver. Further evidence that prostanoids may be involved in the action of arachidonic acid and platelet-activating factor.
The administration of prostaglandin F2 alpha (PGF2 alpha) and the thromboxane A2 analogue, ONO-11113, to rat livers perfused with media containing either 1.3 mM- or 10 microM-Ca2+ was followed by a stimulation of Ca2+ efflux, changes in O2 uptake and glucose output, and increase in portal pressure. The responses elicited by 5 microM-PGF2 alpha were similar to those induced by the alpha-adrenergic agonist phenylephrine. At both 1.3 mM and 10 microM extracellular Ca2+, PGF2 alpha induced Ca2+ efflux (70-90 nmol/g of liver), probably from the same source as that released by phenylephrine. Prostaglandin D2 (5 microM) and prostaglandin E2 (5 microM) also induced responses, but these were generally much smaller (less than 30%) than those induced by PGF2 alpha. Similarly to vasopressin and other Ca2+-mobilizing hormones, PGF2 alpha also interacted synergistically with glucagon (and cyclic AMP) in stimulating Ca2+ influx both in the perfused liver and in isolated hepatocytes. By comparison with phenylephrine and PGF2 alpha, ONO-11113 was much more potent in inducing vasoconstriction, and, at concentrations of 10-200 nM, induced a different pattern of changes in Ca2+ flux, respiration and glycogenolysis. There was first a rapid efflux of Ca2+ (45-60 nmol/g of liver), followed by a smaller Ca2+ influx, and a further release of Ca2+ (approx. 90 nmol/g of liver) when ONO-11113 was removed. Respiration was first stimulated but then markedly inhibited. At concentrations less than 5 nM, ONO-11113 induced a sustained stimulation of O2 uptake and a more prolonged efflux of Ca2+, with less Ca2+ efflux occurring upon the removal of the agent. Glycogenolysis followed a pattern which was similar to the Ca2+ response. Co-administration of glucagon did not potentiate Ca2+ influx by ONO-11113, but the action of ONO-11113 was inhibited (50%) by a few minutes' prior administration of 10 nM-vasopressin. The vasoconstrictive action of ONO-11113 was synergistically potentiated by the co-administration of phenylephrine. Since the actions of arachidonic acid, platelet-activating factor and lysophosphatidylcholine in liver were recently found to be cyclo-oxygenase-sensitive, the results provide strong evidence that at least PGF2 alpha and thromboxane A2 may be involved in mediating the action of these agents.
Animals↗
Non-parenchymal cells as mediators of physiological responses in liver.
Parenchymal cells (hepatocytes) are the sites at which the principal metabolic functions of the liver are located. In the perfused liver, responses (e.g. vasoconstriction and glycogenolysis) to stimulating agents such as zymosan, platelet-activating factor and arachidonic acid, are inhibited by indomethacin and bromophenacyl bromide, inhibitors of cyclo-oxygenase and phospholipase A2, respectively. Since cultured Kupffer and endothelial cells but not hepatocytes, produce eicosanoids, and since eicosanoids and especially prostaglandins induce similar patterns of responses when added directly to the perfused liver, an involvement of these non-parenchymal cells in mediating the above responses is considered likely. We propose that in most situations the responses induced by these stimulating agents are mediated through a combination of pathways that include interaction of the agents directly with hepatocytes or with vasoactive cells (endothelial and/or smooth muscle cells), or interaction of agents initially with non-parenchymal cells to produce and release eicosanoids, which then subsequently interact with hepatocytes or with vasoactive cells.
Animals↗
Second messengers and the regulation of Ca2+ fluxes by Ca2+-mobilizing agonists in rat liver.
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Animals↗
Phosphatidic acid and arachidonic acid each interact synergistically with glucagon to stimulate Ca2+ influx in the perfused rat liver.
The administration of phosphatidic acid to rat livers perfused with media containing either 1.3 mM- or 10 microM-Ca2+ was followed by a stimulation of Ca2+ efflux, O2 uptake and glucose output. The responses elicited by 100 microM-phosphatidic acid were similar to those induced by the alpha-adrenergic agonist phenylephrine. Contrary to suggestions that phosphatidic acid acts like a Ca2+-ionophore, no net influx of Ca2+ was detected until the phosphatidic acid was removed. Sequential infusions of phenylephrine and phosphatidic acid indicate that the two agents release Ca2+ from the same intracellular source. The co-administration of glucagon (or cyclic AMP) and phosphatidic acid, and also of glucagon and arachidonic acid, led to a synergistic stimulation of Ca2+ uptake of the liver, a feature similar to that observed after the co-administration of glucagon and other Ca2+-mobilizing hormones [Altin & Bygrave (1986) Biochem. J. 238, 653-661]. A notable difference, however, is that the synergistic stimulation of Ca2+ uptake induced by the co-administration of glucagon and arachidonic acid was inhibited by indomethacin, whereas that induced by glucagon and phosphatidic acid, or glucagon and other Ca2+-mobilizing agents, was not. The results suggest that the synergistic action of glucagon and arachidonic acid in stimulating Ca2+ influx is mediated by prostanoids, but that of glucagon and phosphatidic acid is evoked by a mechanism similar to that of Ca2+-mobilizing agents.
Animals↗
Evidence that Ca2+ fluxes and respiratory, glycogenolytic and vasoconstrictive effects induced by the action of platelet-activating factor and L-alpha-lysophosphatidylcholine in the perfused rat liver are mediated by products of the cyclo-oxygenase pathway.
The administration of 'acetylglyceryl ether phosphorylcholine' (AGEPC, also known as platelet-activating factor) and L-alpha-lysophosphatidylcholine (LPC) to rat livers perfused with media containing 1.3 mM-Ca2+ was followed by a concentration-dependent efflux of Ca2+ from the liver. Near-maximal response was observed at 100 nM-AGEPC and 50 microM-LPC, and resulted in a net efflux of approx. 130 nmol of Ca2+/g of liver. Onset of Ca2+ efflux occurred about 10 s after AGEPC and LPC administration, reached a maximum after about 50 s (the maximum rate of efflux was approx. 180 nmol/min per g) and thereafter decreased rapidly, and was sometimes followed by a much smaller influx of Ca2+. Sequential infusions of AGEPC or LPC, and phenylephrine, indicate that each of these agents mobilizes Ca2+ from the same intracellular source. The efflux of Ca2+ was not observed in the presence of indomethacin or bromophenacyl bromide, or when the liver was perfused with low-Ca2+-containing (25 microM) media. Other physiological responses, such as changes in respiration, glucose output and portal pressure, were also inhibited under these conditions. The results suggest that the Ca2+-flux changes and other responses are mediated by prostaglandins produced and released within the liver, possibly by cell types other than hepatocytes.
Acetophenones↗