Calcium transport, Ca2(+)-ATPase, and lipid order in rabbit ocular lens membranes

1991 ◽  
Vol 260 (4) ◽  
pp. C731-C737 ◽  
Author(s):  
N. A. Delamere ◽  
C. A. Paterson ◽  
D. Borchman ◽  
K. L. King ◽  
S. A. Cawood
Keyword(s):  
Atpase Activity ◽  
Temperature Dependence ◽  
Calcium Transport ◽  
Calcium Uptake ◽  
Calcium Ionophore ◽  
Membrane Vesicles ◽  
Lipid Phase ◽  
Ionophore A23187 ◽  
Lipid Order ◽  
Uptake Process

Calcium transport was monitored by measuring ATP-dependent 45Ca uptake into membrane vesicles prepared from rabbit lens cortex. Calcium-stimulated adenosinetriphosphatase (Ca2(+)-ATPase) activity was also measured in the same membrane preparation. Both uptake and Ca2(+)-ATPase activity were inhibited by vanadate. Calcium activation of the uptake process was similar to that of the Ca2(+)-ATPase. Calcium uptake was prevented by calcium ionophore A23187, suggesting that the calcium transported into the vesicles remains diffusible. The ATP-dependent calcium uptake probably represents the transport of calcium into “inside-out” membrane vesicles by the Ca2(+)-ATPase mechanism that normally shifts calcium outward from the lens cytoplasm. The temperature dependence of the Ca2(+)-ATPase and the calcium uptake process was determined. Because lipid order can influence Ca2(+)-ATPase function, we attempted to correlate calcium transport with the physical state of the membrane lipids. Infrared spectroscopy was used to determine the temperature dependence of the CH2 symmetric stretching frequency (an order parameter) in the lipids. A similarity was noted between the temperature-dependence curves for lipid order, Ca2(+)-ATPase, and calcium uptake rate. Entropy, enthalpy, and transition temperature calculated for the Ca2(+)-ATPase and calcium uptake process were in the same range as those parameters calculated for the lipid-phase transition.

scholarly journals Characteristics of the Ca2+ pump and Ca2+-ATPase in the plasma membrane of rat myometrium

1988 ◽  
Vol 252 (1) ◽  
pp. 215-220 ◽  
Author(s):  
A Enyedi ◽  
J Minami ◽  
A J Caride ◽  
J T Penniston
Keyword(s):  
Plasma Membrane ◽  
Atpase Activity ◽  
Calcium Uptake ◽  
Limited Proteolysis ◽  
Membrane Vesicles ◽  
Calcium Pump ◽  
Ionophore A23187 ◽  
Plasma Membrane Vesicles ◽  
Active Calcium ◽  
Stimulation Of

A plasma membrane-enriched fraction from rat myometrium shows ATP-Mg2+-dependent active calcium uptake which is independent of the presence of oxalate and is abolished by the Ca2+ ionophore A23187. Ca2+ loaded into vesicles via the ATP-dependent Ca2+ uptake was released by extravesicular Na+. This showed that the Na+/Ca2+ exchange and the Ca2+ uptake were both occurring in plasma membrane vesicles. In a medium containing KCl, vanadate readily inhibited the Ca2+ uptake (K1/2 5 microM); when sucrose replaced KCl, 400 microM-vanadate was required for half inhibition. Only a slight stimulation of the calcium pump by calmodulin was observed in untreated membrane vesicles. Extraction of endogenous calmodulin from the membranes by EGTA decreased the activity and Ca2+ affinity of the calcium pump; both activity and affinity were fully restored by adding back calmodulin or by limited proteolysis. A monoclonal antibody (JA3) directed against the human erythrocyte Ca2+ pump reacted with the 140 kDa Ca2+-pump protein of the myometrial plasma membrane. The Ca2+-ATPase activity of these membranes is not specific for ATP, and is not inhibited by mercurial agents, whereas Ca2+ uptake has the opposite properties. Ca2+-ATPase activity is also over 100 times that of calcium transport; it appears that the ATPase responsible for transport is largely masked by the presence of another Ca2+-ATPase of unknown function. Measurements of total Ca2+-ATPase activity are, therefore, probably not directly relevant to the question of intracellular Ca2+ control.

ATP-dependent calcium transport across basal plasma membranes of human placental trophoblast

1987 ◽  
Vol 252 (1) ◽  
pp. C38-C46 ◽  
Author(s):  
G. J. Fisher ◽  
L. K. Kelley ◽  
C. H. Smith
Keyword(s):  
Plasma Membrane ◽  
Calcium Transport ◽  
Plasma Membranes ◽  
Calcium Uptake ◽  
Regulatory Protein ◽  
Calcium Ionophore ◽  
Maximum Velocity ◽  
Free Calcium ◽  
Ionophore A23187 ◽  
Basal Plasma

As a first step in understanding the cellular basis of maternal-fetal calcium transfer, we examined the characteristics of calcium uptake by a highly purified preparation of the syncytiotrophoblast basal (fetal facing) plasma membrane. In the presence of nanomolar concentrations of free calcium, basal membranes demonstrated substantial ATP-dependent calcium uptake [K0.5 = 119 nM, maximum velocity (Vmax) = 2 nM X min-1 X mg-1]. This uptake required magnesium, was not significantly affected by Na+ or K+ (50 mM), or sodium azide (10 mM). Intravesicular calcium was rapidly and completely released by the calcium ionophore A23187. Calcium transport was significantly stimulated by the calcium-dependent regulatory protein calmodulin. Placental membrane fractions enriched in endoplasmic reticulum (ER) and mitochondria also demonstrated ATP-dependent calcium uptake. In contrast to basal membrane, mitochondrial calcium uptake was completely inhibited by azide. The rate of calcium uptake by the ER was only 20% of that of basal membranes. We conclude that the placental basal plasma membrane possesses a high-affinity calcium transport system similar to that found in plasma membranes of a variety of cell types. This transporter is situated to permit it to function in vivo in maternal-fetal calcium transfer.

Calcium transport by rat duodenal villus and crypt basolateral membranes

1987 ◽  
Vol 252 (2) ◽  
pp. G170-G177 ◽  
Author(s):  
J. R. Walters ◽  
M. M. Weiser
Keyword(s):  
Vitamin D ◽  
Calcium Transport ◽  
Calcium Uptake ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Calcium Pump ◽  
Cell Fraction ◽  
Basolateral Membranes ◽  
Pump Activity ◽  
Crypt Cells

Rat duodenal cells were isolated sequentially to give fractions enriched for villus and crypt cells. From each of these fractions, basolateral-enriched membrane vesicles were prepared and ATP-dependent calcium uptake was studied. Calcium uptake was sensitive to temperature, was inhibited by vanadate and by A23187, and was lower in vitamin D-deficient animals. In normal animals, calcium transport was approximately twofold greater in villus-tip than in crypt cell-fraction basolateral membranes though the affinity of the uptake for calcium was similar (Km = 0.3 microM). In vitamin D-deficient animals, the crypt-to-villus gradient was reduced, and in all fractions, calcium transport was similar to or lower than that in the crypts of normal animals. Six hours after vitamin D-deficient animals were repleted with 1,25-dihydroxycholecalciferol, a significant increase in calcium transport by everted gut sacs was present; however, basolateral calcium transport was significantly increased in only the mid-villus fractions, and no change was seen in the villus-tip fractions. Thus vitamin D appears necessary for the development of increased basolateral membrane calcium pump activity in duodenal villus cells, but not all cells in vitamin D-deficient rats are able to respond to 1,25-dihydroxycholecalciferol.

Mechanism of regulation of calcium-pumping activity in chick intestine

1992 ◽  
Vol 262 (5) ◽  
pp. G797-G805
Author(s):  
J. Takito ◽  
T. Shinki ◽  
H. Tanaka ◽  
T. Suda
Keyword(s):  
Vitamin D ◽  
Calcium Transport ◽  
Calcium Uptake ◽  
Early Stage ◽  
Basolateral Membrane ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Membrane Vesicles ◽  
Intestinal Calcium Transport ◽  
Pumping Activity

The role of the calcium pump in the stimulation of intestinal calcium transport activity by 1 alpha,25-dihydroxyvitamin D3 [1 alpha,25(OH)2D3] was examined in chicks. The in situ intestinal absorption of calcium increased approximately threefold in the duodenum, jejunum, and ileum 6 h after a single injection of 625 ng of 1 alpha,25(OH)2D3 into vitamin D-deficient chicks. The same treatment also increased approximately twofold the rate of ATP-dependent calcium uptake by the basolateral membrane vesicles (BL) isolated from those three sites. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that a Mg(2+)-dependent calcium-stimulated phosphorylated intermediate with an apparent molecular mass of 105 kDa appeared in the BL. The 1 alpha,25(OH)2D3 treatment gave no change in the levels of the intermediate. Pretreatment of the BL with alkaline phosphatase decreased the calcium uptake by the BL isolated from 1 alpha,25(OH)2D3-treated chicks, but it had little effect on the uptake by the BL from vitamin D-deficient chicks. These results suggest that at an early stage of the 1 alpha,25(OH)2D3-induced intestinal calcium transport process, the vitamin regulates the calcium-pumping activity of chick intestinal BL by phosphorylation and dephosphorylation but not by a stoichiometric change in the pump.

Effect of parathyroid hormone on transport by toad and turtle bladder

1987 ◽  
Vol 252 (1) ◽  
pp. R63-R68
Author(s):  
S. Sabatini ◽  
N. A. Kurtzman
Keyword(s):  
Parathyroid Hormone ◽  
Epithelial Cells ◽  
Calcium Uptake ◽  
Short Circuit ◽  
Calcium Ionophore ◽  
Toad Bladder ◽  
Base Line ◽  
Ionophore A23187 ◽  
45Ca Uptake ◽  
Turtle Bladder

We recently demonstrated that parathyroid hormone (PTH) inhibited both vasopressin- and cyclic AMP-stimulated water transport in the toad bladder. This was associated with an increase in calcium uptake by isolated epithelial cells. We postulated that PTH exerts its action on H2O transport by directly stimulating calcium uptake. The current study was designed to compare the effects of PTH and the calcium ionophore, A23187, on H2O and Na transport and H+ secretion in toad and turtle bladders. In toad bladder, PTH and A23187 decreased arginine vasopressin (AVP)-stimulated H2O flow and short-circuit current (SCC) after 60 min serosal incubation. In turtle bladder A23187 decreased SCC to 79.3 +/- 3.6% of base line (P less than 0.05), and significantly decreased RSCC as well. PTH had no effect on SCC or H+ secretion in turtle bladders. Both PTH and A23187 increased 45Ca uptake in toad bladder epithelial cells; only A23187 increased 45Ca uptake in the turtle bladder. The different action of PTH in these two membranes, compared with that of the calcium ionophore, illustrates the selectivity of PTH on membrane transport. PTH increases calcium uptake and decreases transport only in a hormone-sensitive epithelium, whereas the ionophore works in virtually all living membranes. The mode of action of these two agents to increase calcium uptake is, therefore, likely different.

scholarly journals Calcium uptake and release by isolated cortices and microsomes from the unfertilized egg of the sea urchin Strongylocentrotus droebachiensis.

1986 ◽  
Vol 102 (6) ◽  
pp. 2205-2210 ◽  
Author(s):  
J A Oberdorf ◽  
J F Head ◽  
B Kaminer
Keyword(s):  
Sea Urchin ◽  
Calcium Uptake ◽  
Inositol Trisphosphate ◽  
Calcium Ionophore ◽  
Strongylocentrotus Droebachiensis ◽  
Cortical Granule ◽  
Free Calcium ◽  
Cortical Region ◽  
Ionophore A23187 ◽  
Dependent Manner

Isolated cortices from unfertilized sea urchin eggs sequester calcium in an ATP-dependent manner when incubated in a medium containing free calcium levels characteristic of the resting cell (approximately 0.1 microM). This ATP-dependent calcium uptake activity was measured in the presence of 5 mM Na azide to prevent mitochondrial accumulation, was increased by oxalate, and was blocked by 150 microM quercetin and 50 microM vanadate (known inhibitors of calcium uptake into the sarcoplasmic reticulum). Cortical regions preloaded with 45Ca in the presence of ATP were shown to dramatically increase their rate of calcium efflux upon the addition of (a) the calcium ionophore A23187 (10 microM), (b) trifluoperazine (200 microM), (c) concentrations of free calcium that activated cortical granule exocytosis, and (d) the calcium mobilizing agent inositol trisphosphate. This pool of calcium is most likely sequestered in the portion of the egg's endoplasmic reticulum that remains associated with the cortical region during its isolation. We have developed a method for obtaining a high yield of purified microsomal vesicles from whole eggs. This preparation also demonstrates ATP-dependent calcium sequestering activity which increases in the presence of oxalate and has similar sensitivities to calcium transport inhibitors; however, the isolated microsomal vesicles did not show any detectable release of calcium when exposed to inositol trisphosphate.

Peptide models for protein-mediated cation transport

1980 ◽  
Vol 58 (10) ◽  
pp. 865-870 ◽  
Author(s):  
Charles M. Deber
Keyword(s):  
Calcium Transport ◽  
Calcium Ionophore ◽  
Cation Transport ◽  
Benzyl Ester ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Design And Synthesis ◽  
Cellular Processes ◽  
Naturally Occurring ◽  
Peptide Models

Substances which can perturb the transmembrane cation balance in a predictable manner have wide-ranging uses in the study of cellular processes. We have undertaken to examine transmembrane calcium transport on the molecular level through the design and synthesis of a series of ionophoric peptides as models for protein-mediated calcium transport. General mechanisms for carrier-mediated membrane transport are discussed. Cation transport profiles are presented for transport by synthetic peptides of structure cyclo(Glu(OR)-Sar-Gly-(N-R1)-Gly)2, where R = benzyl ester or H; R1 = n-decyl or cyclohexyl. Transport of physiologically abundant cations across "liquid membranes" in Pressman cells mediated by cyclo(Glu-Sar-Gly-(N-decyl)Gly)2 was observed to be essentially calcium specific, as long as calcium ions were present in the system. Multilamellar and unilamellar phosphatidylcholine vesicles were each found to be emptied of internal 45Ca2+ ions upon addition of cyclo(Glu(OBz)-Sar-Gly-(N-cyclohexyl)Gly)2 to the vesicle suspension. The results are compared with the naturally occurring calcium ionophore A23187.

scholarly journals Selective regulation of the Rab9-independent transport of ricin to the Golgi apparatus by calcium

2002 ◽  
Vol 115 (17) ◽  
pp. 3449-3456 ◽  
Author(s):  
Silje U. Lauvrak ◽  
Alicia Llorente ◽  
Tore-Geir Iversen ◽  
Kirsten Sandvig
Keyword(s):  
Golgi Apparatus ◽  
Calcium Transport ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Independent Process ◽  
Mannose 6 Phosphate ◽  
Phosphoinositide 3 Kinase ◽  
Modified Forms

Transport of ricin from endosomes to the Golgi apparatus occurs, in contrast to the transport of the mannose 6-phosphate receptor, by a Rab9-independent process. To characterize the pathway of ricin transport to the Golgi apparatus, we investigated whether it was regulated by calcium. As shown here, our data indicate that calcium is selectively involved in the regulation of ricin transport to the Golgi apparatus. Thapsigargin, which inhibits calcium transport into the ER, and the calcium ionophore A23187 both increased the transport of ricin to the Golgi apparatus by a factor of 20. By contrast, transport of the mannose 6-phosphate receptor to the Golgi apparatus was unaffected. Ricin and mannose 6-phosphate receptor transport were measured by quantifying the sulfation of modified forms of ricin and the mannose 6-phosphate receptor. The increased transport of ricin was reduced by wortmannin and LY294002, suggesting that phosphoinositide 3-kinase might be involved in transport of ricin to the Golgi apparatus. Together, these findings indicate that the different pathways to the Golgi apparatus utilized by ricin and the mannose 6-phosphate receptor are regulated by different mechanisms.

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