scholarly journals Intracellular calcium mobilization and activation of the Na+/H+ exchanger in platelets

1993 ◽  
Vol 290 (2) ◽  
pp. 617-622 ◽  
Author(s):  
E Poch ◽  
A Botey ◽  
J Gaya ◽  
A Cases ◽  
F Rivera ◽  
...  
Keyword(s):  
Intracellular Calcium ◽  
Fluorescent Dyes ◽  
Extracellular Calcium ◽  
Free Calcium ◽  
Regulatory Relationship ◽  
Cytosolic Ph ◽  
Intracellular Calcium Mobilization ◽  
Free Calcium Concentration ◽  
Cytosolic Free Calcium Concentration ◽  
Intracellular Alkalinization

The aim of the present study was to evaluate the regulatory relationship between the cytosolic free calcium concentration ([Ca2+]i and cytosolic pH (pHi). [Ca2+]i and pHi were measured using the fluorescent dyes fura-2 and BCECF [2′,7′-bis-(carboxyethyl)-5,6-carboxyfluorescein] respectively. In a medium with 1 mmol/l extracellular calcium, thrombin (2.5 units/ml) induced an increment in [Ca2+]i of 638 +/- 31 nmol/l (n = 5) and an intracellular alkalinization of 0.14 +/- 0.01 pH units (n = 8). Both responses were dependent on the concentration of thrombin, displaying a sigmoidal dose-response pattern. The intracellular alkalinization was dependent upon extracellular Na+ and was amiloride-sensitive, indicating that it was mediated by activation of the Na+/H+ exchanger. When extracellular calcium was chelated with EGTA prior to the addition of thrombin, the intracellular alkalinization was not affected (0.15 +/- 0.02 at 2.5 units/ml thrombin, n = 8). Under these circumstances, the [Ca2+]i increment represents mobilization from internal stores, reaching 157 +/- 42 nmol/l at 2.5 units/ml thrombin. When platelets were preloaded with the intracellular calcium chelator MAPTAM (1,2-bis-5-methylaminophenoxylethane-NNN'-tetraacetoxymethyl acetate) to block the increase in [Ca2+]i induced by thrombin, no increment in pHi was observed. Moreover, MAPTAM-loaded calcium-depleted platelets had a basal pHi that was more acidic than in the presence of 1 mmol/l extracellular calcium (6.93 +/- 0.09 versus 7.14 +/- 0.01, n = 26, P < 0.001). Ionomycin induced an elevation of [Ca2+]i that was accompanied by a concomitant increase in pHi, which was Na(+)-dependent and amiloride-sensitive. [Ca2+]i and pHi increases induced by ionomycin were both dependent on the concentration of ionomycin. In conclusion, an increase in [Ca2+]i is necessary for the agonist-induced activation of the Na+/H+ exchanger in platelets. Non-agonist-induced increases in [Ca2+]i seems to prompt activation of the exchanger. In addition, Ca(2+)-depleted platelets have a more acidic basal pHi, indicating that the basal level of [Ca2+]i is also important for maintaining the basal pHi.

Cytosolic free calcium in normal somatotropes: effects of forskolin and phorbol ester

1989 ◽  
Vol 256 (3) ◽  
pp. E375-E379
Author(s):  
R. W. Holl ◽  
M. O. Thorner ◽  
D. A. Leong
Keyword(s):  
Growth Hormone ◽  
Phorbol Ester ◽  
Plaque Assay ◽  
Extracellular Calcium ◽  
Free Calcium ◽  
Cytosolic Free Calcium ◽  
Gh Secretion ◽  
Free Calcium Concentration ◽  
Cytosolic Free Calcium Concentration ◽  
A Site

Digital imaging microscopy using the calcium-sensitive indicator probe fura-2 was combined with a reverse hemolytic plaque assay (RHPA) for growth hormone (GH) secretion. This technique allows dynamic measurements of the cytosolic free calcium concentration ([Ca2+]i) in individual pituitary somatotropes. Stimulation by growth hormone-releasing factor (GRF) increases, whereas somatostatin (SRIF) reduces [Ca2+]i in this cell type. [Ca2+]i increased in somatotropes when the cellular content of adenosine 3',5'-cyclic monophosphate (cAMP) was elevated by 1) activating cellular adenylate cyclase with forskolin (5 microM) and 2) treatment with the cAMP-analogues dibutyryl-cAMP (1 mM) or 8-bromo-cAMP (5 mM). The forskolin-induced calcium rise was abolished in the absence of extracellular calcium. This indicates that cAMP increases the influx of calcium into the cytosol and thereby stimulates hormone release. When forskolin was given in combination with SRIF (10 nM), [Ca2+]i decreased to the same level reached with SRIF treatment alone, indicating a site of action distal to the generation of cAMP. Activating protein kinase C with the phorbol ester 12,13-phorbol dibutyrate (PDB; 100 nM) increased [Ca2+]i as well. Again, this effect was dependent on extracellular calcium and blocked when PDB and SRIF were applied simultaneously. Combined stimulation with GRF plus PDB did not augment the response of [Ca2+]i over GRF treatment alone.

V2-like vasopressin receptor mobilizes intracellular Ca2+ in rat medullary collecting tubules

1993 ◽  
Vol 265 (1) ◽  
pp. F35-F45 ◽  
Author(s):  
A. Champigneulle ◽  
E. Siga ◽  
G. Vassent ◽  
M. Imbert-Teboul
Keyword(s):  
Intracellular Calcium ◽  
Calcium Concentration ◽  
Free Calcium ◽  
Cytosolic Free Calcium ◽  
Free Calcium Concentration ◽  
Cytosolic Free Calcium Concentration ◽  
The Mean ◽  
V2 Antagonist ◽  
Collecting Tubules ◽  
Calcium Pool

Cytosolic free calcium concentration ([Ca2+]i) was measured in single microdissected rat medullary collecting tubules [outer (OMCD) and inner (IMCD)] to identify receptors involved in vasopressin (AVP)-induced [Ca2+]i increases. In both segments, [Phe2,Orn8]vasotocin ([Phe2,Orn8]VT), a specific V1 agonist, as well as the V2 agonist 1-desamino-8-D-AVP (dDAVP) triggered [Ca2+]i variations. In OMCD, the mean response to 10 nM AVP roughly corresponded to the sum of V1 and V2 agonists effects. In IMCD, dDAVP (10 nM) alone reproduced the calcium response to AVP (delta[Ca2+]i = 243 +/- 34 nM, n = 6, and 248 +/- 27 nM, n = 8, with dDAVP and AVP, respectively). Furthermore, in the same experiments V1 and V2 maximal effects were not additive ([Phe2,Orn8]VT = 154 +/- 21 nM, n = 6; dDAVP + [Phe2,Orn8]VT = 233 +/- 23 nM, n = 9). As AVP, dDAVP released intracellular calcium (delta[Ca2+]i in calcium-free medium = 182 +/- 24 nM, n = 8, vs. 182 +/- 14 nM, n = 6 with 10 nM dDAVP and AVP, respectively). Neither 8-(4-chlorophenyl-thio)-adenosine 3',5'-cyclic monophosphate nor forskolin modified [Ca2+]i. A cross-reaction of dDAVP with an oxytocin (OT) receptor can be excluded since 1) the specific OT agonist [Thr4,Gly7]OT (10 nM) increased only slightly [Ca2+]i (delta-[Ca2+]i = 20 +/- 5 nM, n = 11); 2) the dDAVP response was not altered by the specific OT antagonist [1-(beta-mercapto-beta,beta-cyclopentamethylene propionic acid),2-(O-methyl)tyrosine,4-threonine, 8-ornithine,9-tyrosylamide]vasotocin [d(CH2)5(1),O-Me-Tyr2,Thr4,Tyr-NH2(9)]OVT; 3) it was insensitive to V1 antagonists but was totally blocked by the V1/V2 antagonist [d(CH2)5(1),O-Et-Tyr2,Val4]AVP ([delta[Ca2+]i = 18 +/- 4 nM, n = 6). These results indicate that in IMCD AVP increases [Ca2+]i via both V1 and V2 receptors. [Ca2+]i variations due to V2 receptors involve a mechanism independent of adenylate cyclase and coupled to the same intracellular calcium pool as V1 and V2 receptors.

Intracellular calcium and phospholipid turnover are not involved in the inhibition of iodothyronine 5'-deiodinase type II activity by T4

1991 ◽  
Vol 124 (1) ◽  
pp. 67-75
Author(s):  
Cristiana Juge-Aubry ◽  
Pierre Dôme ◽  
Catherine A. Siegrist-Kaiser ◽  
Alessandro M. Capponi ◽  
Albert G. Burger
Keyword(s):  
Intracellular Calcium ◽  
Calcium Concentration ◽  
Enzyme Inactivation ◽  
Free Calcium ◽  
Type Ii ◽  
Cytosolic Free Calcium ◽  
Experimental Conditions ◽  
Phospholipid Turnover ◽  
Free Calcium Concentration ◽  
Cytosolic Free Calcium Concentration

Abstract. In glial cell cultures, iodothyronine 5'-deiodinase type II is stimulated by dibutyryl cAMP. Serum-free medium increases enzyme activity and prolongs the half-life of the enzyme. T4 and rT3 specifically inhibit this activity. We tested whether enzyme inactivation by T4 was mediated by changes in cytosolic free calcium concentration and/or phospholipid turnover. Intracellular calcium concentration was decreased either by chelation of extracellular calcium or by chelation of extracellular and intracellular calcium. Neither basal hypothyroid 5'-deiodinase activity nor its inactivation by T4 were modified in such experimental conditions, compared with control cells incubated in normal calcium-containing medium. T4 by itself had no effect on the cytosolic free calcium concentration for up to 20 min. Studies on phospholipid turnover included norepinephrine in parallel to T4 as positive stimulation control. While norepinephrine clearly accelerated phosphoinositide turnover, there was no effect of T4 on any phospholipid turnover. These results suggest that neither cytosolic free calcium nor phospholipid turnover is involved in T4-dependent modulation of 5'-deiodinase type II activity in astrocytes in culture.

scholarly journals The Lectin Chaperone Calnexin Is Involved in the Endoplasmic Reticulum Stress Response by Regulating Ca2+ Homeostasis in Aspergillus nidulans

2017 ◽  
Vol 83 (15) ◽  
Author(s):  
Shenghua Zhang ◽  
Hailin Zheng ◽  
Qiuyi Chen ◽  
Yuan Chen ◽  
Sha Wang ◽  
...  
Keyword(s):  
Er Stress ◽  
Aspergillus Nidulans ◽  
Molecular Chaperone ◽  
Hyphal Growth ◽  
Extracellular Calcium ◽  
Free Calcium ◽  
Content Type ◽  
Free Calcium Concentration ◽  
Cytosolic Free Calcium Concentration ◽  
Limiting Condition

ABSTRACT The Ca2+-mediated signaling pathway is crucial for environmental adaptation in fungi. Here we show that calnexin, a molecular chaperone located in the endoplasmic reticulum (ER), plays an important role in regulating the cytosolic free calcium concentration ([Ca2+]c) in Aspergillus nidulans. Inactivation of calnexin (ClxA) in A. nidulans caused severe defects in hyphal growth and conidiation under ER stress caused by the ER stress-inducing agent dithiothreitol (DTT) or high temperature. Importantly, defects in the ΔclxA mutant were restored by the addition of extracellular calcium. Furthermore, the CchA/MidA complex (the high-affinity Ca2+ channels), calcineurin (calcium/calmodulin-dependent protein phosphatase), and PmrA (secretory pathway Ca2+ ATPase) were required for extracellular calcium-based restoration of the DTT/thermal stress sensitivity in the ΔclxA mutant. Interestingly, the ΔclxA mutant exhibited markedly reduced conidium formation and hyphal growth defects under the low-calcium condition, which is similar to defects caused by mutations in MidA/CchA. Moreover, the phenotypic defects were further exacerbated in the ΔclxA ΔmidA ΔcchA mutant, which suggested that ClxA and MidA/CchA are both required under the calcium-limiting condition. Using the calcium-sensitive photoprotein aequorin to monitor [Ca2+]c in living cells, we found that ClxA and MidA/CchA complex synergistically coordinate transient increase in [Ca2+]c in response to extracellular calcium. Moreover, ClxA, in particular its luminal domain, plays a role in mediating the transient [Ca2+]c in response to DTT-induced ER stress in the absence of extracellular calcium, indicating ClxA may mediate calcium release from internal calcium stores. Our findings provide new insights into the role of calnexin in the regulation of calcium-mediated response in fungal ER stress adaptation. IMPORTANCE Calnexin is a well-known molecular chaperone conserved from yeast to humans. Although it contains calcium binding domains, little is known about the role of calnexin in Ca2+ regulation. In this study, we demonstrate that calnexin (ClxA) in the filamentous fungus Aspergillus nidulans, similar to the high-affinity calcium uptake system (HACS), is required for normal growth and conidiation under the calcium-limiting condition. The ClxA dysfunction decreases the transient cytosolic free calcium concentration ([Ca2+]c) induced by a high extracellular calcium or DTT-induced ER stress. Our findings provide the direct evidence that calnexin plays important roles in regulating Ca2+ homeostasis in addition to its role as a molecular chaperone in fungi. These results provide new insights into the roles of calnexin and expand knowledge of fungal stress adaptation.

Platelet Cytosolic Free Calcium in Essential Hypertension: Responses to Vasopressin

1989 ◽  
Vol 77 (2) ◽  
pp. 183-188 ◽  
Author(s):  
A. F. Dominiczak ◽  
J. J. Morton ◽  
G. Murray ◽  
P. F. Semple
Keyword(s):  
Essential Hypertension ◽  
Arginine Vasopressin ◽  
Calcium Concentration ◽  
Systolic Pressure ◽  
Extracellular Calcium ◽  
Free Calcium ◽  
Cytosolic Free Calcium ◽  
Hypertensive Patients ◽  
Free Calcium Concentration ◽  
The Difference

1. Resting and stimulated free calcium concentrations have been measured in platelets loaded with the fluorescent probe quin2 from 30 patients with essential hypertension and from 30 age-matched controls. 2. Cytosolic free calcium concentrations were 94.6 ± 2.7 (mean ± sem) in the hypertensive group and 91.7 ± 2.8 nmol/l in the normotensive group, the difference was not significant. 3. Arginine vasopressin caused a transient increase in platelet free calcium concentration in all subjects. In the presence of extracellular calcium the increase was significantly higher in the control subjects than in the hypertensive patients (P = 0.005). In the absence of extracellular calcium, arginine vasopressin caused much smaller increases, and there was then no difference between the responses of the two groups. 4. Platelet free calcium concentrations were measured again in 13 patients after 8 weeks treatment with either verapamil (n = 6) or atenolol (n = 7). The reductions in systolic pressure after drug treatment were correlated with the changes in cytosolic free calcium concentrations (r = 0.75, P < 0.01).

Angiotensin II causes sustained elevations in cytosolic calcium in glomerulosa cells

1988 ◽  
Vol 255 (3) ◽  
pp. E338-E346 ◽  
Author(s):  
R. E. Kramer
Keyword(s):  
Angiotensin Ii ◽  
Calcium Concentration ◽  
Cytosolic Calcium ◽  
Calcium Signal ◽  
Free Calcium ◽  
Cytosolic Free Calcium ◽  
Cytosolic Calcium Concentration ◽  
Free Calcium Concentration ◽  
Cytosolic Free Calcium Concentration ◽  
Glomerulosa Cells

Studies were conducted to examine the effects of angiotensin II on cytosolic free calcium concentration in bovine adrenal glomerulosa cells maintained in primary culture. The calcium indicator, fura-2, and discontinuous dual-wavelength fluorescence spectroscopy were used to measure cytosolic free calcium in superfused adherent cell monolayers. Basal cytosolic free calcium concentration was 63.7 +/- 3.3 nM. The threshold concentration for angiotensin II-stimulated increases in cytosolic calcium was 10(-14)-10(-13) M, and maximal elevation of cytosolic calcium was produced by 10(-9) M angiotensin II. Angiotensin II (10(-13) M) produced a gradual increase in cytosolic calcium concentration that plateaued after 3-5 min of superfusion at a level approximately 1.2 times that of control cells. The calcium signal invoked by a maximal concentration (10(-9) M) of angiotensin II, in contrast, was characterized by an immediate, intense (approximately 8-fold) increase in cytosolic calcium concentration that decayed within 5 min to a lower, but sustained, level 2.5-3 times that of control cells. The calcium signals invoked by intermediate concentrations (10(-12)-10(-10) M) of angiotensin II exhibited dose-dependent increases in magnitude and a gradual transition in nature between those invoked by threshold and maximal concentrations of the peptide. The effect of angiotensin II to increase cytosolic calcium concentration was accompanied by an increase in aldosterone output. The increase in steroidogenesis was most closely correlated with the magnitude of the initial calcium signal. At high concentrations (10(-10) and 10(-9) M) of angiotensin II, there was a clear dissociation between aldosterone output and the magnitude of the sustained calcium signal.(ABSTRACT TRUNCATED AT 250 WORDS)

Vasoconstriction: a new activity for platelet-derived growth factor

Science ◽  
1986 ◽  
Vol 232 (4746) ◽  
pp. 87-90 ◽  
Author(s):  
BC Berk ◽  
RW Alexander ◽  
TA Brock ◽  
MA Gimbrone ◽  
RC Webb
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Rat Aorta ◽  
Platelet Derived Growth Factor ◽  
Free Calcium ◽  
Free Calcium Concentration ◽  
Cytosolic Free Calcium Concentration

Platelet-derived growth factor (PDGF) is a potent mitogen for vascular smooth muscle cells that has been implicated in the pathogenesis of atherosclerosis. The potential role of PDGF in the altered vasoreactivity of atherosclerotic vessels has been studied through an examination of its effects on contractility in the rat aorta. PDGF caused a concentration-dependent contraction of aortic strips and was significantly more potent on a molar basis than the classic vasoconstrictor peptide angiotensin II. Furthermore, PDGF increased the cytosolic free calcium concentration in cultured rat aortic smooth muscle cells. These observations suggest a new biological activity for PDGF that may contribute to the enhanced vasoreactivity of certain atherosclerotic vessels.

Sign in / Sign up

Export Citation Format

Share Document