Relationship between hormonal activation of phosphatidylinositol hydrolysis, fluid secretion and calcium flux in the blowfly salivary gland

The addition of 5-hydroxytryptamine to the isolated blowfly salivary gland stimulates fluid secretion, transepithelial calcium transport and the breakdown of 32P- or 3H-labelled phosphatidylinositol The breakdown of [32P]phosphatidylcholine and [32P]-phosphatidylethanolamine was not stimulated by 5-hydroxytryptamine. In salivary glands incubated with myo-[2-3H]inositol for 1–3 h, more than 95% of the label retained by the tissue was in the form of phosphatidylinositol. The addition of 5-hydroxytryptamine resulted in an increase in the accumulation of label in intracellular inositol 1:2-cyclic phosphate, inositol 1-phosphate and free inositol along with an increase in the release of [3H]inositol to the medium and saliva. The release of [3H]inositol to the medium served as a sensitive indicator of phosphatidylinositol breakdown. The release of [3H]inositol was not increased by cyclic AMP or the bivalent-cation ionophore A23187 under conditions in which salivary secretion was accelerated. The stimulation of fluid secretion by low concentrations of 5-hydroxytryptamine was potentiated by 3-isobutyl-1-methylxanthine, which had no effect on inositol release. The stimulation of fluid secretion by 5-hydroxytryptamine was greatly reduced in calcium-free buffer, but the breakdown of phosphatidylinositol continued at the same rate in the absence of calcium. These results support the hypothesis that breakdown of phosphatidylinositol by 5-hydroxytryptamine is involved in the gating of calcium.

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The relationship of calcium to receptor-controlled stimulation of phosphatidylinositol turnover. Effects of acetylcholine, adrenaline, calcium ions, cinchocaine and a bivalent cation ionophore on rat parotid-gland fragments

Biochemical Journal ◽

10.1042/bj1480479 ◽

1975 ◽

Vol 148 (3) ◽

pp. 479-485 ◽

Cited By ~ 65

Author(s):

L M Jones ◽

R H Michell

Keyword(s):

Parotid Gland ◽

De Novo ◽

Head Group ◽

Ionophore A23187 ◽

Bivalent Cation ◽

Phosphatidylinositol Turnover ◽

Rat Parotid Gland ◽

Muscarinic Cholinergic ◽

Rat Parotid ◽

Stimulation Of

The possibility that Ca2+ ions are involved in the control of the increased phosphatidylinositol turnover which is provoked by alpha-adrenergic or muscarinic cholinergic stimulation of rat parotid-gland fragments has been investigated. Both types of stimulation provoked phosphatidylinositol breakdown, which was detected either chemically or radiochemically, and provoked a compensatory synthesis of the lipid, detected as an increased rate of incorporation of 32Pi into phosphatidylinositol. Acetylcholine had little effect on the incorporation of labelled glycerol, whereas adrenaline stimulated it significantly, but to a much lower extent than 32P incorporation: this suggests that the response to acetylcholine was entirely accounted for by renewal of the phosphorylinositol head-group of the lipid, but that some synthesis de novo was involved in the response to adrenaline. The responses to both types of stimulation, whether measured as phosphatidylinositol breakdown or as phosphatidylinositol labelling, occurred equally well in incubation media containing 2.5 mm-Ca2+ or 0.2 mm-EGTA [ethanedioxybis(ethylamine)-tetra-acetic acid]. Incubation with a bivalent cation ionophore (A23187) led to a small and more variable increase in phosphatidylinositol labelling with 32Pi, which occurred whether or not Ca2+ was available in the extracellular medium: this was not accompanied by significant phosphatidylinositol breakdown. Cinchocaine, a local anaesthetic, produced parallel increases in the incorporation of Pi and glycerol into phosphatidylinositol. This is compatible with its known ability to inhibit phosphatidate phosphohydrolase (EC 3.1.3.4) and increase phosphatidylinositol synthesis de novo in other cells. These results indicate that the phosphatidylinositol turnover evoked by alpha-adrenergic or muscarinic cholinergic stimuli in rat parotid gland probably does not depend on an influx of Ca2+ into the cells in response to stimulation. This is in marked contrast with the K+ efflux from this tissue, which is controlled by the same receptors, but is strictly dependent on the presence of extracellular Ca2+. The Ca2+-independence of stimulated phosphatidylinositol metabolism may mean that it is controlled through a mode of receptor function different from that which controls other cell responses. Alternatively, it can be interpreted as indicating that stimulated phosphatidylinositol breakdown is intimately involved in the mechanisms of action of alpha-adrenergic and muscarinic cholinergic receptor systems.

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The alteration of mitotic events by ionophore A23187 and carbonyl cyanide n-chlorophenylhydrazone

Journal of Cell Science ◽

10.1242/jcs.75.1.347 ◽

1985 ◽

Vol 75 (1) ◽

pp. 347-355

Author(s):

M.L. Ziegler ◽

J.E. Sisken ◽

S. Vedbrat

Keyword(s):

Oxidative Phosphorylation ◽

Intracellular Calcium ◽

Calcium Ions ◽

Hela Cells ◽

Cytosolic Calcium ◽

Ionophore A23187 ◽

Chromosome Movement ◽

Low Concentrations ◽

Carbonyl Cyanide ◽

Stimulation Of

A large quantity of published work indicates that calcium ions may be involved in the regulation of mitotic events and recent reports suggest that the onset of chromosome movement is dependent upon a transient increase in free cytosolic calcium ions. In this paper we examine the effects of two agents known to perturb intracellular calcium pools on mitosis in HeLa cells. These were the calcium-selective ionophore A23187 and carbonyl cyanide n-chlorophenylhydrazone (CCCP), which is a protonophoric inhibitor of oxidative phosphorylation. Owing to a stimulation of glycolysis, the latter agent does not decrease intracellular ATP in HeLa but does cause mitochondria to release calcium ions. Our data show that, at low concentrations, both agents prolong metaphase but differ in their effects on anaphase and cytokinesis. Studies with chlorotetracycline, a commonly used probe for membrane-associated calcium, verify that these agents do affect calcium pools under the conditions of our experiments. The data presented are consistent with the idea that increased cytosolic calcium levels can directly or indirectly affect mitotic events but, contrary to other suggestions, cause a prolongation of metaphase, i.e. they delay the onset of chromosome movement.

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Forskolin as an activator of cyclic AMP accumulation and secretion in blowfly salivary glands

Biochemical Journal ◽

10.1042/bj2040147 ◽

1982 ◽

Vol 204 (1) ◽

pp. 147-151 ◽

Cited By ~ 19

Author(s):

I Litosch ◽

Y Saito ◽

J N Fain

Keyword(s):

Salivary Gland ◽

Cyclic Amp ◽

Adenylate Cyclase ◽

Salivary Glands ◽

Mammalian Cells ◽

Salivary Secretion ◽

Gland Secretion ◽

Cyclic Amp Accumulation ◽

Salivary Gland Secretion ◽

Stimulation Of

Forskolin is a diterpene that activates adenylate cyclase in a variety of mammalian cells. In addition of forskolin to blowfly salivary glands increased cyclic AMP accumulation and salivary secretion. There was a small increase in transepithelial movement of labelled Ca2+. Forskolin did not induce breakdown of labelled phosphatidylinositol or inhibit the stimulation of phosphatidylinositol breakdown caused by 5-hydroxytryptamine. These data indicate that forskolin can mimic all the effects of 5-hydroxytryptamine on salivary-gland secretion that have been attributed to cyclic AMP.

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Inhibition of phosphatidylinositol synthesis and the inactivation of calcium entry after prolonged exposure of the blowfly salivary gland to 5-hydroxytryptamine

Biochemical Journal ◽

10.1042/bj1780059 ◽

1979 ◽

Vol 178 (1) ◽

pp. 59-69 ◽

Cited By ~ 129

Author(s):

M J Berridge ◽

J N Fain

Keyword(s):

Salivary Gland ◽

Phosphatidic Acid ◽

Salivary Glands ◽

Calcium Transport ◽

Prolonged Exposure ◽

Calcium Entry ◽

Ionophore A23187 ◽

Hydrolysis Of ◽

External Calcium ◽

Stimulation Of

The incorporation of [32P]Pi into all salivary-gland phospholipids except phosphatidic acid was inhibited by 5-hydroxytryptamine. The accumulation of [32P]Pi into phosphatidic acid was actually enhanced by 5-hydroxytryptamine. There was an inhibition of labelled inositol incorporation into phosphatidylinositol by 5-hydroxytryptamine, which seems to be mediated by calcium because it was mimicked by the ionophore A23187, but was prevented if glands were stimulated with 5-hydroxytryptamine in the absence of external calcium. Inhibition of synthesis together with stimulation of breakdown will decrease the concentration of phosphatidylinositol, which could account for the inactivation of calcium transport observed at high 5-hydroxytryptamine concentrations. When salivary glands were stimulated with 1 micrometer-5-hydroxytryptamine, there was a rapid increase in the transfer of 45Ca2+ from the medium into the saliva, but with time this transport declined to a low value. If the glands were washed free of 5-hydroxytryptamine and incubated in the presence of 2mM-inositol for 1 h, the increase in calcium transport caused by 5-hydroxytryptamine was restored. There was little recovery in the absence of inositol. If glands were stimulated with 5-hydroxytryptamine in the absence of external calcium, a condition which prevents the inhibition of phosphatidylinositol synthesis, calcium transport in response to 5-hydroxytryptamine was greater than in glands preincubated with 5-hydroxytryptamine in the presence of calcium. The inactivation of calcium transport may result from a decrease in phosphatidylinositol concentration. These results support the hypothesis that the hydrolysis of phosphatidylinositol plays some role in either the opening or closing of calcium ‘gates’.

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The characteristics of intracellular Ca2+ signals in vivo necessitate a new model for salivary fluid secretion

10.1101/2020.12.30.424864 ◽

2021 ◽

Author(s):

Takahiro Takano ◽

Amanda M. Wahl ◽

Kai-Ting Huang ◽

John Rugis ◽

James Sneyd ◽

...

Keyword(s):

Mathematical Model ◽

Fluid Secretion ◽

Salivary Secretion ◽

Apical Region ◽

Isolated Cells ◽

Marked Contrast ◽

K Channels ◽

Stimulation Of

AbstractSalivary fluid secretion involves an intricate choreography to result in the trans-epithelial movement of NaCl and water into the acinus lumen. Current models are based on experimental observations in enzymatically isolated cells where the Ca2+ signal invariably propagates globally and thus appears ideally suited to activate spatially separated Cl and K channels. We monitored Ca2+ signals and salivary secretion in live mice expressing GCamp6F, following stimulation of the nerves innervating the submandibular gland. Consistent with in vitro studies, Ca2+ signals were initiated in the apical endoplasmic reticulum. In marked contrast to in vitro data, highly localized trains of Ca2+ transients that failed to propagate from the apical region were observed. Following stimuli optimum for secretion, large apical-basal gradients were elicited. Given this incompatibility to the previous model, a new mathematical model was constructed to explain how salivary secretion can be efficiently stimulated by apically localized Ca2+ signals.

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Highly Localized intracellular Ca2+ signals promote optimal salivary gland fluid secretion

eLife ◽

10.7554/elife.66170 ◽

2021 ◽

Vol 10 ◽

Author(s):

Takahiro Takano ◽

Amanda Wahl ◽

Kai-Ting Huang ◽

Takanori Narita ◽

John Rugis ◽

...

Keyword(s):

Fluid Secretion ◽

Salivary Secretion ◽

Membrane Transporters ◽

Apical Region ◽

Isolated Cells ◽

Marked Contrast ◽

K Channels ◽

Basolateral Plasma Membrane ◽

Stimulation Of

Salivary fluid secretion involves an intricate choreography of membrane transporters to result in the trans-epithelial movement of NaCl and water into the acinus lumen. Current models are largely based on experimental observations in enzymatically isolated cells where the Ca2+ signal invariably propagates globally and thus appears ideally suited to activate spatially separated Cl and K channels, present on the apical and basolateral plasma membrane, respectively. We monitored Ca2+ signals and salivary secretion in live mice expressing GCamp6F, following stimulation of the nerves innervating the submandibular gland. Consistent with in vitro studies, Ca2+ signals were initiated in the apical endoplasmic reticulum. In marked contrast to in vitro data, highly localized trains of Ca2+ transients that failed to fully propagate from the apical region were observed. Following stimuli optimum for secretion, large apical-basal gradients were elicited. A new mathematical model, incorporating these data was constructed to probe how salivary secretion can be optimally stimulated by apical Ca2+ signals.

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Calcium ions modulate hormonally stimulated progesterone production in isolated ovarian cells

Biochemical Journal ◽

10.1042/bj2020381 ◽

1982 ◽

Vol 202 (2) ◽

pp. 381-386 ◽

Cited By ~ 28

Author(s):

J D Veldhuis ◽

P A Klase

Keyword(s):

Cyclic Amp ◽

Calcium Ions ◽

Ionophore A23187 ◽

Bivalent Cation ◽

Progesterone Production ◽

Ovarian Cells ◽

Deficient Medium ◽

Time Periods ◽

Short Time ◽

Stimulation Of

Swine granulosa-luteal cells incubated in Ca2+-deficient medium (5 muM final Ca2+ concentration) for short time periods produced diminished quantities of progesterone in response to lutropin. Maximally stimulating effects of prostaglandin E2 and L-adrenaline were also impaired significantly. Diminished progesterone production could not be attributed to alterations in protein synthesis or cell viability. Under Ca2+-deprived conditions, the stimulatory actions of cholera toxin, 3-isobutyl-1-methylxanthine and 8-bromo cyclic AMP were also significantly impeded. Administration of a presumptive antagonist of transmembrane Ca2+ influx (verapamil) or of EGTA to chelate extracellular Ca2+, significantly decreased the total cellular content of Ca2+, and antagonized the actions of lutropin. Micromolar concentrations of trifluoperazine mimicked the suppressive effects of Ca2+ deprivation. Conversely, the bivalent-cation ionophore, ionophore A23187, significantly augmented the stimulation of progesterone produced by lutropin. Thus the present observations implicate Ca2+ in the modulation of hormonally stimulated progesterone production in isolated ovarian cells, and suggest that Ca2+ may influence one or more processes distal to, or independent of, cyclic AMP generation. In addition, the susceptibility of progesterone biosynthesis to inhibition by trifluoperazine suggests a possible role for calmodulin in the ovary.

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Actions of some transmitters and their antagonists on salivary secretion in a tick

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1978.235.1.r76 ◽

1978 ◽

Vol 235 (1) ◽

pp. R76-R81 ◽

Cited By ~ 2

Author(s):

W. R. Kaufman

Keyword(s):

Salivary Gland ◽

Fluid Secretion ◽

Ixodid Tick ◽

Salivary Secretion ◽

Cholinergic Receptors ◽

Secretory Response ◽

Amblyomma Hebraeum ◽

Sensory Pathway

Cholinomimetics (pilocarpine, carbachol, physostigmine, acetylcholine, acetyl-beta-methylcholine) and sympathomimetics (dopamine, epinephrine), when injected into the hemolymph, provoked salivary fluid secretion in the female ixodid tick Amblyomma hebraeum Koch. Atropine, but not tubocurarine or toxiferine, abolished pilocarpine-induced secretion without reducing the response to dopamine. Reserpine and guanethidine likewise selectively attenuated pilocarpine-induced secretion. Following extirpation of the synganglion, pilocarpine no longer provoked a secretory response whereas dopamine did. Thus, the salivary gland appears to be innervated directly by catecholaminergic rather than cholinergic secretory nerves. It is suggested that pilocarpine elicits salivation by interacting with muscarinic-type cholinergic receptors situated either on the cell bodies of the secretory nerves, or alternatively in the integrative or sensory pathway.

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The effect of intracellular calcium ions on adrenaline-stimulated adenosine 3′:5′-cyclic monophosphate concentrations in pigeon erythrocytes, studied by using the ionophore A23187

Biochemical Journal ◽

10.1042/bj1580211 ◽

1976 ◽

Vol 158 (2) ◽

pp. 211-221 ◽

Cited By ~ 25

Author(s):

A K Campbell ◽

K Siddle

Keyword(s):

Cyclic Amp ◽

Adenylate Cyclase ◽

Time Course ◽

Ionophore A23187 ◽

Atp Content ◽

Bivalent Cation ◽

Intact Cells ◽

Maximum Inhibition ◽

Partial Reversal ◽

Stimulation Of

1. The bivalent cation ionophore A23187 was used to increase the intracellular concentration of Ca2+ in pigeon erythrocytes to investigate whether the increase in cyclic AMP content caused by adrenaline might be influenced by a change in intracellular Ca2+ in intact cells. 2. Incubation of cells with adrenaline, in the concentration range 0.55--55 muM, resulted in an increase in the concentration of cyclic AMP over a period of 60 min. The effect of adrenaline was inhibited by more than 90% with ionophore A23187 (1.9 muM) in the presence of 1 mM-Ca2+. This inhibition could be decreased by decreasing either the concentration of the ionophore or the concentration of extracellular Ca2+, and was independent of the concentration of adrenaline. 3. The effect of ionophore A23187 depended on the time of incubation. Time-course studies showed that maximum inhibition by ionophore A23187 was only observed when the cells were incubated with the ionophore for at least 15 min before the addition of adrenaline. 4. The inhibition by ionophore A23187 depended on the concentration of extracellular Ca2+. In the absence of Mg2+, ionophore A23187 (1.9 muM) inhibited the effect of adrenaline by approx. 30% without added Ca2+, by approx. 66% with 10 muM-Ca2+ and by more than 90% with concentrations of added Ca2+ greater than 30 muM. However, even in the presence of EGTA [ethanedioxybis(ethylamine)tetra-acetate](0.1--10 mM), ionophore A23187 caused an inhibition of the cyclic AMP response of at least 30%, which may have been due to a decrease in cell Mg2+ concentration. 5. The addition of EGTA after incubation of cells with ionophore A23187 resulted in a partial reversal of the inhibition of the effect of adrenaline. 6. Inclusion of Mg2+ (2 mM) in the incubation medium antagonized the inhibitory action of ionophore A23187. This effect was most marked when the ionophore A23187 was added to medium containing Mg2+ before the addition of the cells. 7. The cellular content of Mg2+ was decreased by approx. 50% after 20 min incubation with ionophore A23187 (1.9 muM) in the presence of Ca2+ (1 mM) but no Mg2+. When Mg2+ (2 mM) was also present in the medium, ionophore A23187 caused an increase of approx. 80% in cell Mg2+ content. Ionophore A23187 had no significant effect on cell K+ content. 8. Ionophore A23187 caused a decrease in cell ATP content under some conditions. Since effects on cyclic AMP content could also be shown when ATP was not significanlty lowered, it appeared that a decrease in ATP in the cells could not explain the effect of ionophore A23187 on cyclic AMP. 9. Ionophore A23187 (1.9 muM), with 1 mM-Ca2+, did not enhance cyclic AMP degradation in intact cells, suggesting that the effect of ionophore A23187 on cyclic AMP content was mediated through an inhibition of adenylate cyclase rather than a stimulation of cyclic AMP phosphodiesterase. 10. It was concluded that in intact pigeon erythrocytes adenylate cyclase may be inhibited by intracellular concentrations of Ca2+ in the range 1-10 muM.

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α1-Adrenergic stimulation of Ca2+ mobilization without phosphorylase activation in hepatocytes from phosphorylase b kinase-deficient gsd/gsd rats

Biochemical Journal ◽

10.1042/bj1980379 ◽

1981 ◽

Vol 198 (2) ◽

pp. 379-383 ◽

Cited By ~ 21

Author(s):

P F Blackmore ◽

J H Exton

Keyword(s):

Strong Evidence ◽

Liver Cells ◽

Ionophore A23187 ◽

Adrenergic Stimulation ◽

Bivalent Cation ◽

Site Of Action ◽

Adrenergic Activation ◽

Stimulation Of

Phenylephrine, vasopressin and the bivalent cation ionophore A23187 mobilized Ca2+ normally, but failed to activate phosphorylase, in hepatocytes from gsd/gsd rats with a deficiency of liver phosphorylase b kinase. These data provide strong evidence that phosphorylase b kinase is the site of action of the Ca2+ mobilized intracellularly during alpha 1-adrenergic activation of phosphorylase in liver cells.

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