Effect of intracellular pH on acetylcholine-induced Ca2+ waves in mouse pancreatic acinar cells

1998 ◽  
Vol 275 (3) ◽  
pp. C810-C817 ◽  
Author(s):  
Antonio González ◽  
Fatima Pfeiffer ◽  
Andreas Schmid ◽  
Irene Schulz
Keyword(s):  
Intracellular Ph ◽  
Acinar Cells ◽  
Propagation Rate ◽  
Spreading Speed ◽  
Pancreatic Acinar Cells ◽  
Acidic Ph ◽  
Basolateral Side ◽  
Inositol 1,4,5 Trisphosphate ◽  
The Relationship ◽  
Cytosolic Acidification

We have used fluo 3-loaded mouse pancreatic acinar cells to investigate the relationship between Ca2+ mobilization and intracellular pH (pHi). The Ca2+-mobilizing agonist ACh (500 nM) induced a Ca2+ release in the luminal cell pole followed by spreading of the Ca2+ signal toward the basolateral side with a mean speed of 16.1 ± 0.3 μm/s. In the presence of an acidic pHi, achieved by blockade of the Na+/H+exchanger or by incubation of the cells in a Na+-free buffer, a slower spreading of ACh-evoked Ca2+ waves was observed (7.2 ± 0.6 μm/s and 7.5 ± 0.3 μm/s, respectively). The effects of cytosolic acidification on the propagation rate of ACh-evoked Ca2+ waves were largely reversible and were not dependent on the presence of extracellular Ca2+. A reduction in the spreading speed of Ca2+ waves could also be observed by inhibition of the vacuolar H+-ATPase with bafilomycin A1 (11.1 ± 0.6 μm/s), which did not lead to cytosolic acidification. In contrast, inhibition of the endoplasmic reticulum Ca2+-ATPase by 2,5-di- tert-butylhydroquinone led to faster spreading of the ACh-evoked Ca2+ signals (25.6 ± 1.8 μm/s), which was also reduced by cytosolic acidification or treatment of the cells with bafilomycin A1. Cytosolic alkalinization had no effect on the spreading speed of the Ca2+ signals. The data suggest that the propagation rate of ACh-induced Ca2+ waves is decreased by inhibition of Ca2+ release from intracellular stores due to cytosolic acidification or to Ca2+ pool alkalinization and/or to a decrease in the proton gradient directed from the inositol 1,4,5-trisphosphate-sensitive Ca2+ pool to the cytosol.

CCK independently activates intracellular trypsinogen and NF-κB in rat pancreatic acinar cells

2001 ◽  
Vol 280 (3) ◽  
pp. C465-C472 ◽  
Author(s):  
Bing Han ◽  
Baoan Ji ◽  
Craig D. Logsdon
Keyword(s):  
Chinese Hamster ◽  
Acinar Cells ◽  
Nuclear Factor Κb ◽  
Pancreatic Acinar Cells ◽  
Pyrrolidine Dithiocarbamate ◽  
Trypsinogen Activation ◽  
Independent Events ◽  
Dna Binding Assay ◽  
The Relationship ◽  
Stimulation Of

In the cholecystokinin (CCK) hyperstimulation model of acute pancreatitis, two early intracellular events, activation of trypsinogen and activation of nuclear factor-κB (NF-κB), are thought to be important in the development of the disease. In this study, the relationship between these two events was investigated. NF-κB activity was monitored by using a DNA binding assay and mob-1 chemokine gene expression. Intracellular trypsin activity was measured by using a fluorogenic substrate. Protease inhibitors including FUT-175, Pefabloc, and E-64d prevented CCK stimulation of intracellular trypsinogen and NF-κB activation. Likewise, the NF-κB inhibitors pyrrolidine dithiocarbamate and N-acetyl-l-cysteine inhibited CCK stimulation of NF-κB and intracellular trypsinogen activation. These results suggested a possible codependency of these two events. However, CCK stimulated NF-κB activation in Chinese hamster ovary-CCKAcells, which do not express trypsinogen, indicating that trypsin is not necessary for CCK activation of NF-κB. Furthermore, adenovirus-mediated expression in acinar cells of active p65 subunits to stimulate NF-κB, or of inhibitory κB-α molecules to inhibit NF-κB, did not affect either basal or CCK-mediated trypsinogen activation. Thus trypsinogen and NF-κB activation are independent events stimulated by CCK.

Transport and interaction of nitrogen oxides and NO2 with CO2-HCO3- transporters in pancreatic acini

1994 ◽  
Vol 267 (2) ◽  
pp. C385-C393 ◽  
Author(s):  
H. Zhao ◽  
X. Xu ◽  
K. Ujiie ◽  
R. A. Star ◽  
S. Muallem
Keyword(s):  
Nitric Oxide ◽  
Nitrogen Oxides ◽  
Acinar Cells ◽  
Cellular Proteins ◽  
Pancreatic Acinar Cells ◽  
Disulfonic Acid ◽  
No Production ◽  
Pancreatic Acini ◽  
Cytosolic Acidification ◽  
Stimulation Of

Recently, we showed that NO2- increases gap junction (GJ) permeability and synchronizes intracellular Ca2+ concentration oscillations in pancreatic acini (Loessburg et al., J. Biol. Chem. 268: 19769-19775, 1993). NO2- is also an end product of nitric oxide (NO) production and metabolism. Because of the effect of NO2- on GJ permeability and the possible importance of NO2- in NO metabolism and cytotoxicity, we used pancreatic acinar cells and intracellular pH (pHi) measurements to study the interaction of nitrogen oxides and NO2- with cellular proteins. Exposing cells to NO2- resulted in a concentration-dependent cytosolic acidification. The acidification did not require the transport of NO2- and was not mediated by diffusion of HNO2. Because the acidification was prevented by CO2-HCO3- and inhibition of carbonic anhydrase, it is possible that other nitrogen oxides present in a solution containing NO2- enter the cells by diffusion and interact with OH- or H2O to stably acidify the cytosol. NO2- itself is shown to be transported by the HCO3- transporters present in the plasma membrane. Thus manipulation of the cellular Cl- gradient and 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) were used to show Cl-/NO2- exchange, whereas stimulation of external Na(+)-dependent amiloride-insensitive and DIDS-sensitive pHi increase in acidified cells was used to demonstrate a Na(+)-(NO2-)n cotransport. Hence NO2- can be a convenient substitute for HCO3- when studying HCO3- transport in an open system. The studies also show that cellular levels of nitrogen oxides and NO2- can be modulated by the cellular HCO3(-)-buffering system.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Inositol 1,4,5-trisphosphate and inositol 1,3,4-trisphosphate formation in Ca2+-mobilizing-hormone-activated cells

1985 ◽  
Vol 232 (1) ◽  
pp. 237-243 ◽  
Author(s):  
G M Burgess ◽  
J S McKinney ◽  
R F Irvine ◽  
J W Putney
Keyword(s):  
Guinea Pig ◽  
Inositol Trisphosphate ◽  
Acinar Cells ◽  
Temporal Correlation ◽  
Dimethyl Sulphoxide ◽  
Pancreatic Acinar Cells ◽  
Hormone Stimulation ◽  
Inositol 1,4,5 Trisphosphate ◽  
Intracellular Messenger ◽  
Stimulation Of

The inositol trisphosphate liberated on stimulation of guinea-pig hepatocytes, pancreatic acinar cells and dimethyl sulphoxide-differentiated human myelomonocytic HL-60 leukaemia cells is composed of two isomers, the 1,4,5-trisphosphate and the 1,3,4-trisphosphate. Inositol 1,4,5-trisphosphate was released rapidly, with no measurable latency on hormone stimulation, and, consistent with its proposed role as an intracellular messenger for Ca2+ mobilization, there was good temporal correlation between its formation and Ca2+-mediated events in these tissues. There was a definite latency before an increase in the formation of inositol 1,3,4-trisphosphate could be detected. In all of these tissues, however, it formed a substantial proportion of the total inositol trisphosphate by 1 min of stimulation. In guinea-pig hepatocytes, where inositol trisphosphate increases for at least 30 min after hormone application, inositol 1,3,4-trisphosphate made up about 90% of the total inositol trisphosphate by 5-10 min. In pancreatic acinar cells, pretreatment with 20 mM-Li+ caused an increase in hormone-induced inositol trisphosphate accumulation. This increase was accounted for by a rise in inositol 1,3,4-trisphosphate; inositol 1,4,5-trisphosphate was unaffected. This finding is consistent with the observation that Li+ has no effect on Ca2+-mediated responses in these cells. The role, if any, of inositol 1,3,4-trisphosphate in cellular function is unknown.

scholarly journals Ca2+-sensitivity of inositol 1,4,5-trisphosphate-mediated Ca2+ release in permeabilized pancreatic acinar cells

1990 ◽  
Vol 265 (3) ◽  
pp. 681-687 ◽  
Author(s):  
P H G M Willems ◽  
M D De Jong ◽  
J J H H M De Pont ◽  
C H Van Os
Keyword(s):  
Acinar Cells ◽  
Inhibitory Effect ◽  
Feedback Mechanism ◽  
Pancreatic Acinar Cells ◽  
Release Mechanism ◽  
Concomitant Increase ◽  
Permeabilized Cells ◽  
Negative Feedback Mechanism ◽  
Inositol 1,4,5 Trisphosphate ◽  
Energy Dependent

Hormonal and phorbol ester pretreatment of pancreatic acinar cells markedly decreases the Ins(1,4,5)P3-induced release of actively stored Ca2+ [Willems, Van Den Broek, Van Os & De Pont (1989) J. Biol. Chem. 264, 9762-9767]. Inhibition occurred at an ambient free Ca2+ concentration of 0.1 microM, suggesting a receptor-mediated increase in Ca2(+)-sensitivity of the Ins(1,4,5)P3-operated Ca2+ channel. To test this hypothesis, the Ca2(+)-dependence of Ins(1,4,5)P3-induced Ca2+ release was investigated. In the presence of 0.2 microM free Ca2+, permeabilized cells accumulated 0.9 nmol of Ca2+/mg of acinar protein in an energy-dependent pool. Uptake into this pool increased 2.2- and 3.3-fold with 1.0 and 2.0 microM free Ca2+ respectively. At 0.2, 1.0 and 2.0 microM free Ca2+, Ins(1,4,5)P3 maximally released 0.53 (56%), 0.90 (44%) and 0.62 (20%) nmol of Ca2+/mg of acinar protein respectively. Corresponding half-maximal stimulatory Ins(1,4,5)P3 concentrations were calculated to be 0.5, 0.6 and 1.4 microM, suggesting that the affinity of Ins(1,4,5)P3 for its receptor decreases beyond 1.0 microM free Ca2+. The possibility that an inhibitory effect of sub-micromolar Ca2+ is being masked by the concomitant increase in size of the releasable store is excluded, since Ca2+ release from cells loaded in the presence of 0.1 or 0.2 microM free Ca2+ and stimulated at higher ambient free Ca2+ was not inhibited below 1.0 microM free Ca2+. At 2.0 and 10.0 microM free Ca2+, Ca2+, Ca2+ release was inhibited by approx. 30% and 75% respectively. The results presented show that hormonal pretreatment does not lead to an increase in Ca2(+)-sensitivity of the release mechanism. Such an increase in Ca2(+)-sensitivity to sub-micromolar Ca2+ is required to explain sub-micromolar oscillatory changes in cytosolic free Ca2+ by a Ca2(+)-dependent negative-feedback mechanism.

scholarly journals Agonist-dependent Phosphorylation of the Inositol 1,4,5-Trisphosphate Receptor

1999 ◽  
Vol 113 (6) ◽  
pp. 851-872 ◽  
Author(s):  
Andrew P. LeBeau ◽  
David I. Yule ◽  
Guy E. Groblewski ◽  
James Sneyd
Keyword(s):  
Acinar Cells ◽  
Calcium Oscillations ◽  
Pancreatic Acinar Cells ◽  
Ip3 Receptors ◽  
Ip3 Receptor ◽  
Open Probability ◽  
Probability Curve ◽  
Long Period ◽  
Inositol 1,4,5 Trisphosphate ◽  
S Period

The properties of inositol 1,4,5-trisphosphate (IP3)-dependent intracellular calcium oscillations in pancreatic acinar cells depend crucially on the agonist used to stimulate them. Acetylcholine or carbachol (CCh) cause high-frequency (10–12-s period) calcium oscillations that are superimposed on a raised baseline, while cholecystokinin (CCK) causes long-period (>100-s period) baseline spiking. We show that physiological concentrations of CCK induce rapid phosphorylation of the IP3 receptor, which is not true of physiological concentrations of CCh. Based on this and other experimental data, we construct a mathematical model of agonist-specific intracellular calcium oscillations in pancreatic acinar cells. Model simulations agree with previous experimental work on the rates of activation and inactivation of the IP3 receptor by calcium (DuFour, J.-F., I.M. Arias, and T.J. Turner. 1997. J. Biol. Chem. 272:2675–2681), and reproduce both short-period, raised baseline oscillations, and long-period baseline spiking. The steady state open probability curve of the model IP3 receptor is an increasing function of calcium concentration, as found for type-III IP3 receptors by Hagar et al. (Hagar, R.E., A.D. Burgstahler, M.H. Nathanson, and B.E. Ehrlich. 1998. Nature. 396:81–84). We use the model to predict the effect of the removal of external calcium, and this prediction is confirmed experimentally. We also predict that, for type-III IP3 receptors, the steady state open probability curve will shift to lower calcium concentrations as the background IP3 concentration increases. We conclude that the differences between CCh- and CCK-induced calcium oscillations in pancreatic acinar cells can be explained by two principal mechanisms: (a) CCK causes more phosphorylation of the IP3 receptor than does CCh, and the phosphorylated receptor cannot pass calcium current; and (b) the rate of calcium ATPase pumping and the rate of calcium influx from the outside the cell are greater in the presence of CCh than in the presence of CCK.

Calcium release by cholecystokinin analogue OPE is IP3 dependent in single rat pancreatic acinar cells

1994 ◽  
Vol 267 (1) ◽  
pp. C220-C228 ◽  
Author(s):  
H. Y. Gaisano ◽  
D. Wong ◽  
L. Sheu ◽  
J. K. Foskett
Keyword(s):  
Molecular Weight ◽  
Low Molecular Weight Heparin ◽  
Calcium Release ◽  
Acinar Cells ◽  
Pancreatic Acinar Cells ◽  
Low Molecular Weight ◽  
Ip3 Receptor ◽  
Intact Cells ◽  
Inositol 1,4,5 Trisphosphate ◽  
Low Levels

Cholecystokinin (CCK) and carbachol raise intracellular Ca2+ concentration ([Ca2+]i) in pancreatic acinar cells by elevating inositol 1,4,5-trisphosphate (IP3). CCK analogues JMV-180 and OPE stimulate fully efficacious enzyme secretion and [Ca2+]i oscillations but release Ca2+ from intracellular stores by apparently IP3-independent mechanisms in permeabilized acinar cells. In the present study, we investigated whether OPE mobilizes Ca2+ from IP3-sensitive Ca2+ stores and whether IP3 mediates such responses in single intact cells. OPE and JMV-180 similarly elevated IP3 to low levels compared with those elicited by 10 nM CCK. Depletion of IP3-sensitive stores by elevation of intracellular IP3 using carbachol, microinjection of a nonmetabolizable IP3 analogue, or exposure to thapsigargin, in the absence of extracellular Ca2+, depleted the same Ca2+ stores that were sensitive to OPE. In converse experiments, OPE depleted carbachol- or thapsigargin-sensitive stores, indicating that carbachol-, thapsigargin-, IP3-, and OPE-sensitive Ca2+ stores overlap completely and that stores mobilized by OPE are IP3 sensitive. To determine whether IP3 mediates responses to OPE, cells were microinjected with low-molecular-weight heparin, a competitive inhibited the rise of [Ca2+]i in response to carbachol, OPE, or JMV-180, whereas de-N-sulfated heparin, an inactive heparin, was without effect. These results indicate that CCK analogues release Ca2+ from IP3-sensitive Ca2+ stores by mechanisms involving the IP3 receptor.

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