Thapsigargin defines roles of Ca2+ in initial, sustained, and potentiated stimulation of pepsinogen secretion

1994 ◽  
Vol 266 (4) ◽  
pp. G613-G623 ◽  
Author(s):  
Y. Kitsukawa ◽  
C. Felley ◽  
D. C. Metz ◽  
R. T. Jensen
Keyword(s):  
Guinea Pig ◽  
Initial Phase ◽  
Time Course ◽  
Initial Increase ◽  
Cholecystokinin Octapeptide ◽  
Chief Cells ◽  
Pepsinogen Secretion ◽  
Inositol 1,4,5 Trisphosphate ◽  
Stimulation Of ◽  
Sustained Increase

The roles of Ca2+ in agonist-induced pepsinogen secretion from guinea pig chief cells remain unclear. We used cholecystokinin octapeptide (CCK-8) or secretin alone or with thapsigargin (TG) to clarify these roles. TG releases Ca2+ from intracellular stores by inhibiting microsomal Ca(2+)-adenosinetriphosphatase (ATPase), thereby depleting intracellular Ca2+ (Cai2+) stores. In most cells TG also causes Ca2+ influx. In the present study, with an extracellular Ca2+ concentration ([Ca2+]o) of 1.5 mM, CCK-8 (0.1 microM) caused a rapid increase in pepsinogen secretion; however, the rate decreased with time. With [Ca2+]o = 0, the initial increase was similar but later secretion was abolished, suggesting that Ca2+ influx was important for sustained secretion. With [Ca2+]o = 1.5 mM, TG (0.1 microM) caused a 2.7-fold sustained increase in in Cai2+ concentration ([Ca2+]i) and a ninefold sustained increase in pepsinogen secretion. With [Ca2+]o = 0, TG caused a transient 66% increase in [Ca2+]i and a 50% increase in pepsinogen secretion. The time course of TG-induced pepsinogen secretion correlated with the time course of TG-induced increases in [Ca2+]i. These data demonstrated that Ca2+ influx itself was a potent stimulant of pepsinogen secretion. We further focused on the roles of increasing [Ca2+]i from Cai2+ stores. With or without extracellular Ca2+ (Cao2+) present, addition of CCK-8 (0.1 microM) 10 min after TG caused no further increase in [Ca2+]i, demonstrating depletion of the inositol 1,4,5-trisphosphate-sensitive pool. The Ca(2+)-mobilizing agent CCK-8 caused no pepsinogen secretion 10 min after TG preincubation, demonstrating that mobilization of Ca2+ from intracellular stores was important in the rapid initial phase stimulation of pepsinogen secretion caused by CCK-8. In contrast, preincubation with TG had no effect on pepsinogen secretion by secretin, an agent that increases adenosine 3',5'-cyclic monophosphate. A 6-min preincubation with TG potentiated the subsequent stimulation of pepsinogen secretion caused by secretin in the presence of Cao2+ where [Ca2+]i remained elevated. However, TG-induced potentiations of secretin-stimulated pepsinogen secretion was abolished once [Ca2+]i had returned to the basal level in the absence of Cao2+.(ABSTRACT TRUNCATED AT 400 WORDS)

Functionally distinct receptors for cholecystokinin and gastrin on dispersed chief cells from guinea pig stomach

1988 ◽  
Vol 254 (2) ◽  
pp. G151-G155 ◽  
Author(s):  
J. A. Cherner ◽  
V. E. Sutliff ◽  
D. M. Grybowski ◽  
R. T. Jensen ◽  
J. D. Gardner
Keyword(s):  
Amino Acids ◽  
Guinea Pig ◽  
Receptor Antagonists ◽  
Chief Cells ◽  
Pepsinogen Secretion ◽  
Guinea Pig Stomach ◽  
Cck Receptor Antagonists ◽  
Stimulation Of

Caerulein, gastrin, and C-terminal fragments of cholecystokinin (CCK) varying in length from eight (CCK-8) to four (CCK-4) amino acids stimulate pepsinogen secretion from dispersed chief cells prepared from guinea pig stomach. C-terminal fragments of CCK containing fewer than four amino acids, even when tested at concentrations as high as 3 mM, do not stimulate pepsinogen secretion. The efficacies of gastrin and the various CCK-related peptides, coupled with the pattern of action of CCK receptor antagonists, indicate that chief cells from guinea pig stomach possess two functionally distinct classes of receptors, C-receptors and G-receptors. The C-receptors can be occupied by caerulein, CCK-8, CCK-7, des(SO3)CCK-8, or des(SO3)CCK-7, and occupation of C-receptors causes full stimulation of pepsinogen secretion. G-receptors can be occupied by gastrin I, gastrin II, CCK-6, CCK-5, or CCK-4, and occupation of G-receptors causes stimulation of pepsinogen secretion that is 60% of maximal.

Stimulation of pepsinogen secretion in permeable isolated gastric glands

1985 ◽  
Vol 249 (3) ◽  
pp. G408-G415
Author(s):  
S. H. Norris ◽  
S. J. Hersey
Keyword(s):  
Lactate Dehydrogenase ◽  
Gastric Glands ◽  
Cholecystokinin Octapeptide ◽  
Chief Cells ◽  
Isolated Gastric Glands ◽  
Pepsinogen Secretion ◽  
Calcium Stimulation ◽  
Intracellular Regulation ◽  
Stimulus Secretion Coupling ◽  
Stimulation Of

Rabbit isolated gastric glands were treated with digitonin so that stimulation of pepsinogen secretion could be studied in a permeable system. Criteria for permeabilization were the release of lactate dehydrogenase in response to digitonin as well as the finding that calcium stimulation and spermine inhibition required the presence of digitonin. Other evidence confirmed that digitonin directly permeabilized chief cells. Pepsinogen secretion was elicited from digitonin-treated gastric glands by a number of agents, including calcium, vanadate, cholecystokinin octapeptide (CCK-OP), 8-bromo-adenosine 3',5'-cyclic monophosphate, and forskolin. Spermine was found to inhibit secretion stimulated by each of these agents only in the presence of digitonin, suggesting an intracellular site of spermine action. We concluded that spermine inhibition of secretion could be used as a marker of secretion elicited from permeable chief cells. The ability to stimulate pepsinogen secretion by such agents as CCK-OP and forskolin suggests that stimulus-secretion coupling is virtually intact even in permeable chief cells. We felt that this preparation should offer unusual opportunities for investigating the mechanisms involved in the intracellular regulation and activation of pepsinogen secretion.

scholarly journals Mediation of pepsinogen secretion from guinea pig chief cells by Ca2+/calmodulin-dependent protein kinase II

1995 ◽  
Vol 1268 (2) ◽  
pp. 123-129 ◽  
Author(s):  
Naotsuka Okayama ◽  
Makoto Itoh ◽  
Takashi Joh ◽  
Tadahisa Miyamoto ◽  
Toshihiko Takeuchi ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Guinea Pig ◽  
Dependent Protein Kinase ◽  
Chief Cells ◽  
Pepsinogen Secretion ◽  
Protein Kinase Ii ◽  
Dependent Protein

Prostaglandin E2 desensitizes cAMP-mediated pepsinogen secretion in chief cells

1991 ◽  
Vol 261 (5) ◽  
pp. G858-G865 ◽  
Author(s):  
S. Fiorucci ◽  
K. E. McArthur
Keyword(s):  
Adenylate Cyclase ◽  
Guinea Pig ◽  
Prostaglandin E2 ◽  
Vasoactive Intestinal Peptide ◽  
Adenylate Cyclase System ◽  
Chief Cells ◽  
Cyclic Monophosphate ◽  
Camp Generation ◽  
Pepsinogen Secretion ◽  
In Cells

To evaluate whether pretreatment with prostaglandin E2 (PGE2) could desensitize pepsinogen secretion in chief cells from guinea pig, chief cells were pretreated with 10 microM PGE2 for up to 30 min. Desensitization of subsequent PGE2-stimulated secretion was maximal after 15 min, averaging only 29 +/- 9% (SE) of pepsinogen secretion in control cells stimulated with 10 microM PGE2. Desensitization was half-maximal with 30 nM PGE2. PGE2 pretreatment at 4 degrees C did not cause desensitization. In cells pretreated with 10 microM PGE2 for 15 min and then given 60 min to recover, responsiveness increased to 79 +/- 7% of that for control cells stimulated with PGE2. Thus the desensitization was reversible. Pretreatment with PGD2 and PGF2a did not alter subsequent PGE2-mediated secretion. PGE2-induced desensitization was heterologous but mediator specific because pepsinogen secretion was reduced in response to adenosine 3',5'-cyclic monophosphate (cAMP)-mediated agents (secretin and vasoactive intestinal peptide) but not Ca(2+)-mediated agents (CCK-8, gastrin, or carbachol). Pretreating chief cells with 10 microM PGE2 did not significantly alter cAMP generation in response to PGE2, secretin, or 3-isobutyl-1-methylxanthine, suggesting that desensitization was not mediated by an alteration in the receptor-coupled adenylate cyclase system. Because PGE2 pretreatment also desensitized pepsinogen secretion induced by the synthetic cAMP analogues 8-BrcAMP and 2'-O-monobutyryl-8-BrcAMP, it is likely that the ability of PGE2 to desensitize pepsinogen secretion in chief cells isolated from guinea pig is due to a mechanism distal to generation of cAMP.

Regulation of pepsinogen release from canine chief cells in primary monolayer culture

1983 ◽  
Vol 245 (5) ◽  
pp. G641-G646 ◽  
Author(s):  
M. J. Sanders ◽  
D. A. Amirian ◽  
A. Ayalon ◽  
A. H. Soll
Keyword(s):  
Phosphodiesterase Inhibitor ◽  
Functional Differentiation ◽  
Maximal Response ◽  
Chief Cells ◽  
Epithelial Monolayers ◽  
Pepsinogen Secretion ◽  
Primary Monolayer ◽  
Dose Dependent ◽  
Short Term Culture ◽  
Stimulation Of

To study the regulation of pepsinogen secretion by chief cells, we have developed techniques for the isolation, enrichment, and short-term culture of chief cells from canine stomach. The fundic mucosa was enzyme dispersed and chief cells were enriched to a content of about 70% using an elutriator rotor. After 36 h in culture confluent monolayers formed that were highly enriched in chief cells. Carbachol induced a time-dependent release of pepsinogen into the medium, with about a threefold increase in pepsinogen secretion over controls found after 60 min of incubation. Carbachol stimulation of pepsinogen secretion was dose dependent, with 5 microM producing 50% of the maximal response found at a carbachol concentration of 100 microM. Atropine (100 microM) produced a rightward shift of the dose-response curve, indicating the presence of a muscarinic receptor. Dibutyryl cAMP, 8-bromo-cAMP, and forskolin also markedly stimulated pepsinogen secretion. Secretin and vasoactive intestinal peptide (VIP) stimulated pepsinogen secretion, but the response were of smaller magnitude than found with carbachol or the cAMP analogues. The phosphodiesterase inhibitor isobutylmethylxanthine also caused a small stimulation of pepsinogen secretion but did not enhance the response to secretin or VIP. These findings indicate that epithelial monolayers can spontaneously form from isolated canine chief cells and retain functional differentiation evident by a response to stimulation. Canine chief cells in culture possess muscarinic and secretin receptors and respond to cAMP.

Y2 receptors for peptide YY and neuropeptide Y on dispersed chief cells from guinea pig stomach

1992 ◽  
Vol 262 (4) ◽  
pp. G756-G762
Author(s):  
G. Singh ◽  
L. Singh ◽  
J. P. Raufman
Keyword(s):  
Guinea Pig ◽  
Neuropeptide Y ◽  
Peptide Yy ◽  
Camp Production ◽  
Chief Cells ◽  
High Affinity ◽  
Cell Internalization ◽  
Pepsinogen Secretion ◽  
Guinea Pig Stomach ◽  
Dissociation Constant Kd

Peptide YY (PYY) and neuropeptide Y (NPY) inhibit agonist-induced adenosine 3',5'-cyclic monophosphate (cAMP) production and pepsinogen secretion from chief cells. We used radiolabeled PYY and NPY to characterize receptors on chief cells from guinea pig stomach. Binding of 125I-labeled PYY was rapid (70% maximal within 10 min) and specific (not inhibited by secretin, vasoactive intestinal peptide, cholecystokinin, carbachol, prostaglandin E2, forskolin, or cholera toxin). Measurement of the ability of PYY to inhibit binding of 125I-PYY indicated the presence of 1.8 x 10(3) high-affinity [dissociation constant (Kd) = 1.7 nM] and 5.1 x 10(4) low-affinity (Kd = 83.3 nM) sites/cell. Internalization of bound 125I-PYY was suggested by slow and incomplete dissociation in the presence of unlabeled PYY (50% after 2 h) and was examined further by measuring residual binding after washing with acetic acid (pH 2.5), glycine (pH 10.5), or trypsin. After 30 min at 37 degrees C, internalization of radioligand was evidenced by the failure of washing with these solutions to remove 50-65% of bound radioactivity. At 4 degrees C, internalization of 125I-PYY was nearly abolished. Binding of 125I-PYY and 125I-NPY was inhibited by NPY-(13-36) but not by [Leu31,Pro34]NPY indicating that these are Y2 receptors. In guinea pig chief cells, PYY and NPY modulate cAMP-mediated pepsinogen secretion by interacting with specific high-affinity Y2 receptors.

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.

Synergism between ouabain and monensin in neurogenic responses of guinea-pig vas deferens

1988 ◽  
Vol 66 (5) ◽  
pp. 643-647 ◽  
Author(s):  
Takeshi Katsuragi ◽  
Lulu Kuratomi ◽  
Koji Miyamoto ◽  
Tatsuo Furukawa
Keyword(s):  
Guinea Pig ◽  
Time Course ◽  
Vas Deferens ◽  
Contractile Activity ◽  
Atpase Inhibitor ◽  
Exchange System ◽  
Deficient Medium ◽  
Small Contraction ◽  
Monensin A ◽  
Stimulation Of

Interrelations between ouabain, a Na+–K+ ATPase inhibitor, and monensin, a Na+ ionophore, on noradrenaline liberation and contractile activity were evaluated in the guinea-pig vas deferens. Monensin (1 μM) per se elicited a small contraction of the tissue. However, amplitude and time to the peak of large and sustained contractions evoked by 10 μM ouabain were potentiated and markedly shortened, respectively, by monensin. Contractions elicited by ouabain with or without monensin were prevented by 3 μM phentolamine or by pretreatment with reserpine. Contractions evoked by K+-free solution were augmented by monensin. In an HPLC study, noradrenaline outflow from the vas deferens was moderately and considerably increased by monensin (10 μM) and ouabain (100 μM), respectively. The ouabain-evoked output of noradrenaline was enhanced in the presence of monensin and the time course for maximum noradrenaline release was shortened, as was the contractile activity. This enhanced outflow after ouabain plus monensin was reserpine sensitive but not tetrodotoxin sensitive. Furthermore, this noradrenaline outflow was roughly halved in Na+-deficient medium, but was unaltered in Ca2+-free medium. These findings suggest that the synergistic effect of ouabain and monensin on noradrenaline liberation from the guinea-pig vas deferens may be due to an elevation of cytoplasmic Ca2+ concentrations, presumably resulting from a stimulation of intracellular Na+–Ca2+ exchange system, but not enhanced Ca2+ entry.

Ability of prostaglandin to reduce ethanol injury to dispersed chief cells from guinea pig stomach

1989 ◽  
Vol 256 (4) ◽  
pp. G704-G714 ◽  
Author(s):  
J. A. Cherner ◽  
L. Naik ◽  
A. Tarnawski ◽  
T. Brzozowski ◽  
J. Stachura ◽  
...  
Keyword(s):  
Lactate Dehydrogenase ◽  
Guinea Pig ◽  
Cell Injury ◽  
Protective Action ◽  
Control Cell ◽  
Ethanol Exposure ◽  
Fast Green ◽  
Chief Cells ◽  
Pepsinogen Secretion ◽  
Guinea Pig Stomach

To determine whether prostaglandin exerts a direct action on individual gastric epithelial cells that protects them from ethanol-induced injury, dispersed chief cells from guinea pig stomach were pretreated with 16,16-dimethyl-prostaglandin E2 (dmPGE2) or placebo before incubation with ethanol or control. Cell injury was assessed in terms of exclusion of Fast Green dye, release of lactate dehydrogenase, alterations of ultrastructure, and pepsinogen secretion stimulated by a variety of secretagogues. Of chief cells 60 +/- 2% were stained by Fast Green if incubated with 10% ethanol for 1 h after pretreatment with placebo, whereas only 38 +/- 1% of cells showed Fast Green staining when pretreated with 2.6 microM dmPGE2 before ethanol exposure. Similarly, 63 +/- 2% of cellular lactate dehydrogenase was released from chief cells pretreated with placebo compared with 36 +/- 4% of lactate dehydrogenase released from cells pretreated with 2.6 microM dmPGE2 (P less than 0.01). The prostaglandin's protective effect persisted throughout a 6-h incubation with ethanol. Scanning and transmission electron micrographs demonstrated disintegration of chief cells pretreated with placebo before ethanol exposure, whereas ultrastructural architecture was relatively preserved among chief cells pretreated with dmPGE2. Preincubation with 8 or 10% ethanol inhibited the subsequent stimulation of pepsinogen secretion caused by carbachol, cholecystokinin, A23187, 12-O-tetradecanoylphorbol 13-acetate, forskolin, or 8-bromoadenosine 3',5'-cyclic monophosphate. Pretreatment with dmPGE2 did not reduce the ethanol-induced inhibition of secretion stimulated by any of these secretagogues. These data indicate that dmPGE2 significantly reduces ethanol-induced damage to dispersed chief cells in terms of alterations of membrane permeability and ultrastructure but does not prevent the ethanol-induced impairment of pepsinogen secretion. These findings provide evidence that dmPGE2 exerts a direct but limited protective action on the gastric chief cell, independent of vascular, paracrine, or neural actions.

Actions of peptide YY and neuropeptide Y on chief cells from guinea pig stomach

1991 ◽  
Vol 260 (6) ◽  
pp. G820-G826
Author(s):  
J. P. Raufman ◽  
L. Singh
Keyword(s):  
Guinea Pig ◽  
Neuropeptide Y ◽  
G Proteins ◽  
Peptide Yy ◽  
Chief Cell ◽  
Inhibitory Effects ◽  
Camp Content ◽  
Chief Cells ◽  
Pepsinogen Secretion ◽  
Guinea Pig Stomach

Peptide YY (PYY), found in intestinal endocrine cells, and neuropeptide Y (NPY), a structural analogue of PYY found in neurons, inhibit gastric, pancreatic, and intestinal fluid and electrolyte secretion. We examined the effects of these peptides on dispersed chief cells from guinea pig stomach. PYY and NPY, but not pancreatic polypeptide, starting at nanomolar concentrations, caused a 40–50% inhibition of secretin-, vasoactive intestinal polypeptide-, prostaglandin E2-, and forskolin-induced increases in chief cell adenosine 3',5'-cyclic monophosphate (cAMP) content and pepsinogen secretion. These inhibitory peptides did not alter pepsinogen secretion caused by cholecystokinin, carbamylcholine, A23187, 8-bromo-cAMP, or a phorbol ester. The inhibitory effects of PYY on chief cell cAMP production occurred within 30 s, were independent of phosphodiesterase activity, and did not affect the actions of cholera toxin. However, the inhibitory effects of PYY were abolished when chief cells were preincubated with pertussis toxin, an agent that uncouples inhibitory guanine nucleotide binding (G) proteins from their receptors. In gastric chief cells, PYY and NPY attenuate the stimulatory effects of secretagogues whose actions are mediated by changes in cellular levels of cAMP. PYY-induced attenuation of chief cell adenylate cyclase activity appears to involve activation of inhibitory G proteins.

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