scholarly journals Phospholipase C-γ1 is required for the activation of store-operated Ca2+ channels in liver cells

2007 ◽  
Vol 405 (2) ◽  
pp. 269-276 ◽  
Author(s):  
Tom Litjens ◽  
Than Nguyen ◽  
Joel Castro ◽  
Edoardo C. Aromataris ◽  
Lynette Jones ◽  
...  
Keyword(s):  
Phospholipase C ◽  
Rat Hepatocytes ◽  
Cell Types ◽  
Liver Cells ◽  
Maximal Effect ◽  
Experimental Conditions ◽  
Induced Changes ◽  
Molecular Components ◽  
Interfering Rna ◽  
Ca2 Channels

Repetitive hormone-induced changes in concentration of free cytoplasmic Ca2+ in hepatocytes require Ca2+ entry through receptor-activated Ca2+ channels and SOCs (store-operated Ca2+ channels). SOCs are activated by a decrease in Ca2+ concentration in the intracellular Ca2+ stores, but the molecular components and mechanisms are not well understood. Some studies with other cell types suggest that PLC-γ (phospholipase C-γ) is involved in the activation of receptor-activated Ca2+ channels and/or SOCs, independently of PLC-γ-mediated generation of IP3 (inositol 1,4,5-trisphosphate). The nature of the Ca2+ channels regulated by PLC-γ has not been defined clearly. The aim of the present study was to determine if PLC-γ is required for the activation of SOCs in liver cells. Transfection of H4IIE cells derived from rat hepatocytes with siRNA (short interfering RNA) targeted to PLC-γ1 caused a reduction (by approx. 70%) in the PLC-γ1 protein expression, with maximal effect at 72–96 h. This was associated with a decrease (by approx. 60%) in the amplitude of the ISOC (store-operated Ca2+ current) developed in response to intracellular perfusion with either IP3 or thapsigargin. Knockdown of STIM1 (stromal interaction molecule type 1) by siRNA also resulted in a significant reduction (approx. 80% at 72 h post-transfection) of the ISOC amplitude. Immunoprecipitation of PLC-γ1 and STIM1, however, suggested that under the experimental conditions these proteins do not interact with each other. It is concluded that the PLC-γ1 protein, independently of IP3 generation and STIM1, is required to couple endoplasmic reticulum Ca2+ release to the activation of SOCs in the plasma membrane of H4IIE liver cells.

scholarly journals Arachidonic acid inhibits the store-operated Ca2+ current in rat liver cells

2005 ◽  
Vol 385 (2) ◽  
pp. 551-556 ◽  
Author(s):  
Grigori Y. RYCHKOV ◽  
Tom LITJENS ◽  
Michael L. ROBERTS ◽  
Greg J. BARRITT
Keyword(s):  
Arachidonic Acid ◽  
Rat Liver ◽  
Membrane Conductance ◽  
Rat Hepatocytes ◽  
Cell Types ◽  
Receptor Activation ◽  
Liver Cells ◽  
H4iie Cells ◽  
Rat Liver Cells ◽  
Ca2 Channels

Vasopressin and other phospholipase-C-coupled hormones induce oscillations (waves) of [Ca2+]cyt (cytoplasmic Ca2+ concentration) in liver cells. Maintenance of these oscillations requires replenishment of Ca2+ in intracellular stores through Ca2+ inflow across the plasma membrane. While this may be achieved by SOCs (store-operated Ca2+ channels), some studies in other cell types indicate that it is dependent on AA (arachidonic acid)-activated Ca2+ channels. We studied the effects of AA on membrane conductance of rat liver cells using whole-cell patch clamping. We found no evidence that concentrations of AA in the physiological range could activate Ca2+-permeable channels in either H4IIE liver cells or rat hepatocytes. However, AA (1–10 μM) did inhibit (IC50=2.4±0.1 μM) Ca2+ inflow through SOCs (ISOC) initiated by intracellular application of Ins(1,4,5)P3 in H4IIE cells. Pre-incubation with AA did not inhibit ISOC development, but decreased maximal amplitude of the current. Iso-tetrandrine, widely used to inhibit receptor-activation of phospholipase A2, and therefore AA release, inhibited ISOC directly in H4IIE cells. It is concluded that (i) in rat liver cells, AA does not activate an AA-regulated Ca2+-permeable channel, but does inhibit SOCs, and (ii) iso-tetrandrine and tetrandrine are effective blockers of CRAC (Ca2+-release-activated Ca2+) channel-like SOCs. These results indicate that AA-activated Ca2+-permeable channels do not contribute to hormone-induced increases or oscillations in [Ca2+]cyt in liver cells. However, AA may be a physiological modulator of Ca2+ inflow in these cells.

Evidence that 2-aminoethyl diphenylborate is a novel inhibitor of store-operated Ca2+ channels in liver cells, and acts through a mechanism which does not involve inositol trisphosphate receptors

2001 ◽  
Vol 354 (2) ◽  
pp. 285-290 ◽  
Author(s):  
Roland B. GREGORY ◽  
Grigori RYCHKOV ◽  
Greg J. BARRITT
Keyword(s):  
Hepatoma Cell ◽  
Regulatory Protein ◽  
Rat Hepatocytes ◽  
Cell Types ◽  
Liver Cells ◽  
Detectable Effect ◽  
Hepatoma Cell Line ◽  
Isolated Rat Hepatocytes ◽  
Fura 2 ◽  
Ca2 Channels

The compound 2-aminoethyl diphenylborate (2-APB), an inhibitor of Ins(1,4,5)P3 receptor action in some cell types, has been used to assess the role of Ins(1,4,5)P3 receptors in the activation of store-operated Ca2+ channels (SOCs) [Ma, Patterson, van Rossum, Birnbaumer, Mikoshiba and Gill (2000) Science 287, 1647–1651]. In freshly-isolated rat hepatocytes, 2-APB inhibited thapsigargin- and vasopressin-stimulated Ca2+ inflow (measured using fura-2) with no detectable effect on the release of Ca2+ from intracellular stores. The concentration of 2-APB which gave half-maximal inhibition of Ca2+ inflow was approx. 10µM. 2-APB also inhibited Ca2+ inflow initiated by a low concentration of adenophostin A but had no effect on maitotoxin-stimulated Ca2+ inflow through non-selective cation channels. The onset of the inhibitory effect of 2-APB on thapsigargin-stimulated Ca2+ inflow was rapid. When 2-APB was added to rat hepatocytes in the presence of extracellular Ca2+ after a vasopressin-induced plateau in the cytoplasmic free Ca2+ concentration ([Ca2+]cyt) had been established, the kinetics of the decrease in [Ca2+]cyt were identical with those induced by the addition of 50µM Gd3+ (gadolinium). 2-APB did not inhibit the release of Ca2+ from intracellular stores induced by the addition of Ins(1,4,5)P3 to permeabilized hepatocytes. In the H4-IIE rat hepatoma cell line, 2-APB inhibited thapsigargin-stimulated Ca2+ inflow (measured using fura-2) and, in whole-cell patch-clamp experiments, the Ins(1,4,5)P3-induced inward current carried by Ca2+. It was concluded that, in liver cells, 2-APB inhibited SOCs through a mechanism which involved the binding of 2-APB to either the channel protein or an associated regulatory protein. 2-APB appeared to be a novel inhibitor of SOCs in liver cells with a mechanism of action which, in this cell type, is unlikely to involve an interaction of 2-APB with Ins(1,4,5)P3 receptors. The need for caution in the use of 2-APB as a probe for the involvement of Ins(1,4,5)P3 receptors in the activation of SOCs in other cell types is briefly discussed.

scholarly journals Effects of taurolithocholate, a Ca2+-mobilizing agent, on cell Ca2+ in rat hepatocytes, human platelets and neuroblastoma NG108-15 cell line

1991 ◽  
Vol 273 (1) ◽  
pp. 153-160 ◽  
Author(s):  
J F Coquil ◽  
B Berthon ◽  
N Chomiki ◽  
L Combettes ◽  
P Jourdon ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Bile Acid ◽  
Cell Line ◽  
Rat Liver ◽  
Neuronal Cell ◽  
Rat Hepatocytes ◽  
Cell Types ◽  
Liver Cells ◽  
Human Platelets ◽  
Rat Liver Cells

The monohydroxy bile acid taurolithocholate permeabilizes the endoplasmic reticulum to Ca2+ in rat liver cells. To assess whether this action on the endoplasmic reticulum was restricted to this tissue, the effects of bile acid were investigated in two cell types quite unrelated to rat hepatocyte, namely human platelets and neuronal NG108-15 cell line. The results showed that taurolithocholate (3-100 microM) had no effect on free cytosolic [Ca2+] in human platelets and NG108-15 cells. whereas it increased it from 180 to 520 nM in rat hepatocytes. In contrast, in cells permeabilized by saponin, taurolithocholate initiated a profound release of the stored Ca2+ from the internal Ca2+ pools in the three cell types. The bile acid released 90% of the Ca2+ pools, with rate constants of about 5 min-1 and half-maximal effects at 15-30 microM. The results also showed that, in contrast with liver cells, which displayed an influx of [14C]taurolithocholate of 2 nmol/min per mg, human platelets and the neuronal cell line appeared to be resistant to [14C]taurolithocholate uptake. The influx measured in these latter cells was about 100-fold lower than in rat liver cells. Taken together, these data suggest that human platelets and NG108-15 cells do not possess the transport system for concentrating monohydroxy bile acids into cells. However, they show that human platelets and neuronal NG108-15 possess, in common with liver cells, the intracellular system responsible for taurolithocholate-mediated Ca2+ release from internal stores.

scholarly journals Histones and basic polypeptides activate Ca2+/cation influx in various cell types

1998 ◽  
Vol 331 (2) ◽  
pp. 623-630 ◽  
Author(s):  
Alessandra GAMBERUCCI ◽  
Rosella FULCERI ◽  
Paola MARCOLONGO ◽  
William F. PRALONG ◽  
Angelo BENEDETTI
Keyword(s):  
Rat Hepatocytes ◽  
Cell Types ◽  
Membrane Depolarization ◽  
Histone H2a ◽  
Specific Manner ◽  
A Cell ◽  
Cell Depolarization ◽  
Influx Pathways ◽  
Basic Polypeptides ◽  
Ca2 Channels

Histone H2A (1–10 µg/ml) added to Ehrlich ascite cell suspensions promoted: (i) Ca2+ influx, but no apparent intracellular Ca2+ mobilization; (ii) plasma-membrane depolarization and Na+ influx in Ca2+-free medium, which were recovered by Ca2+ readmission; (iii) influx of other cations such as Ba2+, Mn2+, choline+ and N-methyl-d-glucamine+, but not of propidium+, ethidium bromide and Trypan Blue. H2A-induced Ca2+ influx and cell depolarization were: (i) blocked by La3+ and Gd3+, but not by various inhibitors of receptor-activated Ca2+-influx pathways/channels; (ii) mimicked by various basic polypeptides, with Mr > 4000; (iii) prevented or reversed by polyanions such as polyglutamate or heparin; (iv) present in other cell types, such as Jurkat, PC12 and Friend erythroleukaemia cells, but virtually absent from rat hepatocytes and thymocytes. We conclude that cationic proteins/polypeptides, by interacting in a cell-specific manner with the cell surface, can activate in those cells putative non-selective Ca2+ channels and membrane depolarization.

scholarly journals Ca2+-induced changes in the secondary structure of a 60 kDa phosphoinositide-specific phospholipase C from bovine brain cytosol

1988 ◽  
Vol 255 (3) ◽  
pp. 807-812 ◽  
Author(s):  
C Herrero ◽  
M E Cornet ◽  
C Lopez ◽  
P G Barreno ◽  
A M Municio ◽  
...  
Keyword(s):  
Secondary Structure ◽  
Phospholipase C ◽  
Alpha Helix ◽  
Cell Types ◽  
Bovine Brain ◽  
Induced Changes ◽  
Brain Cytosol

The purification to homogeneity of a 60 kDa phosphoinositide-specific phospholipase C from bovine brain cytosol is reported here. This enzyme exhibits the same properties, in terms of response to Ca2+, as does the cytosolic activity in a variety of cell types. We show here that Ca2+ does not appear to modulate the binding of the enzyme to the substrate, but induces dramatic changes in its secondary structure. Therefore we suggest that a decrease in the alpha-helix content of this enzyme correlates with its ability to be activated by Ca2+.

Differential regulation of cyclin D1 and cell death by bile acids in primary rat hepatocytes

2007 ◽  
Vol 293 (1) ◽  
pp. G327-G334 ◽  
Author(s):  
Rui E. Castro ◽  
Joana D. Amaral ◽  
Susana Solá ◽  
Betsy T. Kren ◽  
Clifford J. Steer ◽  
...  
Keyword(s):  
Cell Death ◽  
Cyclin D1 ◽  
Bile Acids ◽  
Transcriptional Activation ◽  
Rat Hepatocytes ◽  
Cytochrome C Release ◽  
Primary Rat Hepatocytes ◽  
Dna Microarray Data ◽  
Induced Changes ◽  
Interfering Rna

Ursodeoxycholic (UDCA) and tauroursodeoxycholic (TUDCA) acids modulate apoptosis and regulate cell-cycle effectors, including cyclin D1. In contrast, deoxycholic acid (DCA) induces cell death and cyclin D1. In this study, we explored the role of cyclin D1 in DCA-induced toxicity and further elucidated the antiapoptotic function of UDCA and TUDCA in primary rat hepatocytes. Cells were incubated with DCA and with or without UDCA or TUDCA for 8–30 h. In addition, hepatocytes were transfected with either an adenovirus expressing cyclin D1 or with a cyclin D1 reporter plasmid with or without bile acids. Finally, cells were cotransfected with short interfering RNA targeting p53. Unlike DCA, both UDCA and TUDCA reduced cyclin D1 expression and transcriptional activation, confirming our previous DNA microarray data. Furthermore, UDCA and TUDCA prevented DCA-induced cyclin D1 and cell death. Cyclin D1 overexpression increased DCA-induced Bax translocation, cytochrome c release, and apoptosis. However, UDCA and TUDCA were less efficient at decreasing cyclin D1 levels as well as DCA-induced changes with overexpression. Finally, after p53 silencing, the effects of cyclin D1 overexpression were almost completely abrogated, whereas UDCA and TUDCA cytoprotective potential was reestablished. In conclusion, cyclin D1 is a relevant player in modulating apoptosis by bile acids, in part through a p53-dependent mechanism.

Mechanisms of capacitative calcium entry

2001 ◽  
Vol 114 (12) ◽  
pp. 2223-2229 ◽  
Author(s):  
James W. Putney ◽  
Lisa M. Broad ◽  
Franz-Josef Braun ◽  
Jean-Philippe Lievremont ◽  
Gary St J. Bird
Keyword(s):  
Plasma Membrane ◽  
Phospholipase C ◽  
Binding Sites ◽  
Cell Types ◽  
Store Depletion ◽  
Blocking Effects ◽  
Conformational Coupling ◽  
Phosphatidylinositol 4 ◽  
Filling State ◽  
Ca2 Channels

Capacitative Ca2+ entry involves the regulation of plasma membrane Ca2+ channels by the filling state of intracellular Ca2+ stores in the endoplasmic reticulum (ER). Several theories have been advanced regarding the mechanism by which the stores communicate with the plasma membrane. One such mechanism, supported by recent findings, is conformational coupling: inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) receptors in the ER may sense the fall in Ca2+ levels through Ca2+-binding sites on their lumenal domains, and convey this conformational information directly by physically interacting with Ca2+ channels in the plasma membrane. In support of this idea, in some cell types, store-operated channels in excised membrane patches appear to depend on the presence of both Ins(1,4,5)P3 and Ins(1,4,5)P3 receptors for activity; in addition, inhibitors of Ins(1,4,5)P3 production that either block phospholipase C or inhibit phosphatidylinositol 4-kinase can block capacitative Ca2+ entry. However, the electrophysiological current underlying capacitative Ca2+ entry is not blocked by an Ins(1,4,5)P3 receptor antagonist, and the blocking effects of a phospholipase C inhibitor are not reversed by the intracellular application of Ins(1,4,5)P3. Furthermore, cells whose Ins(1,4,5)P3 receptor genes have been disrupted can nevertheless maintain their capability to activate capacitative Ca2+ entry channels in response to store depletion. A tentative conclusion is that multiple mechanisms for signaling capacitative Ca2+ entry may exist, and involve conformational coupling in some cell types and perhaps a diffusible signal in others.

scholarly journals Noradrenaline, vasopressin and angiotensin increase Ca2+ influx by opening a common pool of Ca2+ channels in isolated rat liver cells

1984 ◽  
Vol 221 (1) ◽  
pp. 121-127 ◽  
Author(s):  
J P Mauger ◽  
J Poggioli ◽  
F Guesdon ◽  
M Claret
Keyword(s):  
Rat Liver ◽  
Channel Model ◽  
Initial Rate ◽  
Liver Cells ◽  
Maximal Effect ◽  
Isolated Rat Liver ◽  
Common Pool ◽  
Rat Liver Cells ◽  
Ca2 Channels ◽  
Isolated Rat

The effects of the Ca2+-mobilizing hormones noradrenaline, vasopressin and angiotensin on the unidirectional influx of Ca2+ were investigated in isolated rat liver cells by measuring the initial rate of 45Ca2+ uptake. The three hormones increased Ca2+ influx, with EC50 values (concentrations giving half-maximal effect) of 0.15 microM, 0.44 nM and 0.8 nM for noradrenaline, vasopressin and angiotensin respectively. The actions of noradrenaline and angiotensin were evident within seconds after their addition to the cells, whereas the increase in Ca2+ influx initiated by vasopressin was slightly delayed (by 5-15s). The activation of Ca2+ influx was maintained as long as the receptor was occupied by the hormone. The measurement of the resting and hormone-stimulated Ca2+ influxes at different external Ca2+ concentrations revealed Michaelis-Menten-type kinetics compatible with a saturable channel model. Noradrenaline, vasopressin and angiotensin increased both Km and Vmax. of Ca2+ influx. It is proposed that the hormones increase the rate of translocation of Ca2+ through a common pool of Ca2+ channels without changing the number of available channels or their affinity for Ca2+.

scholarly journals Protein kinase A regulates the disposition of Ca2+ which enters the cytoplasmic space through store-activated Ca2+ channels in rat hepatocytes by diverting inflowing Ca2+ to mitochondria

1998 ◽  
Vol 330 (3) ◽  
pp. 1179-1187 ◽  
Author(s):  
C. Kekulu FERNANDO ◽  
B. Roland GREGORY ◽  
J. Greg BARRITT
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Regulatory Protein ◽  
Rat Hepatocytes ◽  
Ruthenium Red ◽  
Maximal Effect ◽  
Isolated Rat Hepatocytes ◽  
Carbonyl Cyanide ◽  
Ca2 Channels ◽  
Kinase A

The roles of a trimeric GTP-binding regulatory protein, protein kinase A and mitochondria in the regulation of store-activated (thapsigargin-stimulated) Ca2+ inflow in freshly-isolated rat hepatocytes were investigated. Rates of Ca2+ inflow were estimated by measuring the increase in the fluorescence of intracellular fura-2 following the addition of extracellular Ca2+ (Ca2+o) to cells incubated in the absence of added Ca2+o. Guanosine 5ʹ-[γ-thio]-triphosphate (GTP[S]) and AlF4- inhibited the thapsigargin-stimulated Ca2+o-induced increase in cytoplasmic free Ca2+ concentration ([Ca2+]c) and this inhibition was prevented by the Rp diastereoisomer of adenosine 3ʹ,5ʹ-(cyclic)phosphoro[thioate]. cAMP, forskolin and glucagon (half-maximal effect at 10 nM) mimicked inhibition of the thapsigargin-stimulated Ca2+o-induced increase in [Ca2+]c by GTP[S], but had little effect on thapsigargin-induced release of Ca2+ from intracellular stores. Azide and carbonyl cyanide p-trifluoromethoxyphenylhydrazone inhibited the thapsigargin-stimulated Ca2+o-induced increase in [Ca2+]c in the presence of increased cAMP (induced by glucagon). In contrast, Ruthenium Red markedly enhanced the thapsigargin-stimulated Ca2+o-induced increase in [Ca2+]c in both the presence and absence of increased cAMP (induced by forskolin and dibutyryl cAMP). It is concluded that, in hepatocytes, protein kinase A regulates the disposition of Ca2+, which enters the cytoplasmic space through store-activated Ca2+ channels, by directing some of this Ca2+ to the mitochondria. The idea that caution should be exercised in using observed values of Ca2+o-induced increase in [Ca2+]c as estimates of rates of agonist-stimulated Ca2+ inflow is briefly discussed.

scholarly journals Epidermal growth factor administration decreases liver glycogen and causes mild hyperglycaemia in mice

1996 ◽  
Vol 315 (1) ◽  
pp. 289-293 ◽  
Author(s):  
Montserrat GRAU ◽  
Francesc TEBAR ◽  
Ignasi RAMÍREZ ◽  
Maria SOLEY
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Rat Hepatocytes ◽  
Maximal Effect ◽  
Dependent Manner ◽  
Isolated Cells ◽  
Experimental Conditions ◽  
Epidermal Growth ◽  
Mouse Hepatocytes

Several laboratories report different effects of epidermal growth factor (EGF) on glycogen metabolism in hepatocytes. The discrepancies may be attributed to differences in the experimental conditions. It is therefore important to establish the actual effect of EGF in vivo. Because large physiological variations of EGF concentration in plasma occur in mice, we used this species to address this question. In freshly isolated mouse hepatocytes, EGF increased glycogen degradation in a dose-dependent manner. The maximal effect (36% increase over basal glycogenolysis) was smaller than maximal effects of classical glycogenolytic hormones like adrenaline or glucagon (more than 150% increase over basal). This is in keeping with the smaller effect of EGF on phosphorylase a activity. In contrast with these hormones, EGF did not inhibit glycolysis. Thus these effects of EGF in mouse hepatocytes are similar to those recently described by us in rat hepatocytes [Quintana, Grau, Moreno, Soler, Ramírez and Soley (1995) Biochem. J. 308, 889–894]. When administered to whole animals, EGF increased phosphorylase a activity, decreased the glycogen content in the liver and caused mild hyperglycaemia. Taking together the results obtained for isolated cells and for whole animals, we suggest that the glucosyl residues released from glycogen are used mostly by the liver rather than released to the circulation. This would be different from the action of the classical glycogenolytic hormones, adrenaline and glucagon.

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