Inhibition of active strontium transport from erythrocyte ghosts by internal calcium: Evidence for a specificity controlling site

Summary The inhibition of strontium transport from erythrocyte ghosts by internal calcium was investigated. When active strontium transport was measured in the presence of increasing levels of internal calcium it was found that the inhibition of strontium transport started at an internal calcium level of 0.3mm and was virtually complete when this concentration reached 1.0mm. It was also noted that calcium transport was virtually constant between concentrations of 0.3 and 1.0mm. This experiment indicated that calcium did not inhibit strontium transport by competing for the active site of the transport system. This inhibition was partially reversed by increasing the internal magnesium concentration from 1 to 4mm. A higher level of magnesium at the time of lysis and during incubation enhanced strontium transport. However, the inhibition remained noncompetitive with respect to calcium. Manganese was also found to support calcium and strontium transport. However, it could not reverse the inhibition of strontium transport by internal calcium at any concentration tested. In fact, manganese restored the inhibition of strontium transport by calcium in ghosts that were prepared and incubated in solutions that had high magnesium levels.     

Calcium sensitivity of dicarboxylate transport in cultured proximal tubule cells

Urinary citrate is an important inhibitor of calcium nephrolithiasis and is primarily determined by proximal tubule reabsorption. The major transporter to reabsorb citrate is Na(+)-dicarboxylate cotransporter (NaDC1), which transports dicarboxylates, including the divalent form of citrate. We previously found that opossum kidney (OK) proximal tubule cells variably express either divalent or trivalent citrate transport, depending on extracellular calcium. The present studies were performed to delineate the mechanism of the effect of calcium on citrate and succinate transport in these cells. Transport was measured using isotope uptake assays. In some studies, NaDC1 transport was studied in Xenopus oocytes, expressing either the rabbit or opossum ortholog. In the OK cell culture model, lowering extracellular calcium increased both citrate and succinate transport by more than twofold; the effect was specific in that glucose transport was not altered. Citrate and succinate were found to reciprocally inhibit transport at low extracellular calcium (<60 μM), but not at normal calcium (1.2 mM); this mutual inhibition is consistent with dicarboxylate transport. The inhibition varied progressively at intermediate levels of extracellular calcium. In addition to changing the relative magnitude and interaction of citrate and succinate transport, decreasing calcium also increased the affinity of the transport process for various other dicarboxylates. Also, the affinity for succinate, at low concentrations of substrate, was increased by calcium removal. In contrast, in oocytes expressing NaDC1, calcium did not have a similar effect on transport, indicating that NaDC1 could not likely account for the findings in OK cells. In summary, extracellular calcium regulates constitutive citrate and succinate transport in OK proximal tubule cells, probably via a novel transport process that is not NaDC1. The calcium effect on citrate transport parallels in vivo studies that demonstrate the regulation of urinary citrate excretion with urinary calcium excretion, a process that may be important in decreasing urinary calcium stone formation.     

Investigation of the Accompaniment of Calcium During Active Calcium Transport from Human Erythrocyte Ghosts

To determine whether a cell metabolite was involved in active calcium transport, the cell contents of human erythrocytes were subjected to high dilutions and the resultant ghosts were checked for their ability to actively transport calcium. It was found that the diluted erythrocyte ghosts did retain their capacity to actively transport calcium and that the characteristics of this transport process appeared to be unaltered by the high dilutions. Calcium analysis of the cell membrane and cell supernatant indicated that almost all of the calcium was lost from the cell solution rather than the cell membrane as active calcium transport proceeded. Therefore it appeared that calcium was able to cross the cell membrane without the aid of a cell metabolite. Investigations with layered erythrocytes indicated that the active transport of calcium was not assisted by centrifugation. Neither inorganic phosphate, pyrophosphate, nor an adenine nucleotide appeared to accompany calcium across the membrane as indicated by total phosphate and inorganic phosphate analysis and 260-nm readings of the deproteinized supernatant.     

Comparison of the Ca-Mg ATPase and calcium transport in rat and human erythrocytes: Evidence for an electrogenic mechanism

Inside out vesicles prepared from rat erythrocytes transport calcium by an electrogenic mechanism. Calmodulin stimulates the activity of a Ca-Mg ATPase, and also stimulates calcium transport. Permeant anions stimulate calcium transport relative to that observed when an impermeant anion (gluconate) is employed. The inside out vesicles transport phosphate anion in a calmodulin-stimulated, calcium- and ATP-dependent manner. Membrane potential sensitive fluorescent dyes demonstrate a positive interior membrane potential during calcium transport in the absence of permeant anions. The properties of the rat erythrocyte calcium transport system corresponds closely to those of the human erythrocyte; the activity of the Ca-Mg ATPase, rates and stoichiometries of calcium and phosphate transport and the degree of calmodulin stimulation are directly comparable between the two species.     

Chronic metabolic acidosis stimulated transcellular and solvent drag-induced calcium transport in the duodenum of female rats

Chronic metabolic acidosis results in a negative calcium balance as a result of bone resorption and renal calcium loss. However, reports on the changes in intestinal calcium transport have been controversial. The present investigation therefore aimed to study the effects of chronic metabolic acidosis induced by 1.5% NH(4)Cl administration on the three components of duodenal calcium transport, namely, solvent drag-induced, transcellular active, and passive paracellular components, in rats using an in vitro Ussing chamber technique. The relative mRNA expression of genes related to duodenal calcium transport was also determined. We found that 21-day chronic metabolic acidosis stimulated solvent drag-induced and transcellular active duodenal calcium transport but not passive paracellular calcium transport. Our results further demonstrated that an acute direct exposure to serosal acidic pH, in contrast, decreased solvent drag-induced calcium transport in a pH-dependent fashion but had no effect on transcellular active calcium transport. Neither the transepithelial resistance nor duodenal permeability to Na(+), Cl(-), and Ca(2+) via the passive paracellular pathway were altered by chronic metabolic acidosis, suggesting that widening of the tight junction and changes in the charge-selective property of the tight junction did not occur. Thus the enhanced duodenal calcium transport observed in chronic metabolic acidosis could have resulted from a long-term adaptation, possibly at the molecular level. RT-PCR study revealed that chronic metabolic acidosis significantly increased the relative mRNA expression of duodenal genes associated with solvent drag-induced transport, i.e., the beta(1)-subunit of Na(+)-K(+)-ATPase, zonula occludens-1, occludin, and claudin-3, and with transcellular active transport, i.e., transient receptor potential vanilloid family Ca(2+) channels 5 and 6 and plasma membrane Ca(2+)-ATPase isoform 1b. Total plasma calcium and free ionized calcium and magnesium concentrations were also increased, whereas serum parathyroid hormone and 1alpha,25-dihydroxyvitamin D(3) levels were not changed. The results indicated that 21-day chronic metabolic acidosis affected the calcium metabolism in rats partly through enhancing the mRNA expression of crucial duodenal genes involved in calcium absorption, thereby stimulating solvent drag-induced and transcellular active calcium transport in the duodenum.     

Stimulation of calcium transport of sarcoplasmic reticulum vesicles by the calcium complex of ethylene glycol bis(beta-aminoethyl ether)-N,N',-tetraacetic acid

The calcium ion dependence of calcium transport by isolated sarcoplasmic reticulum vesicles from rabbit skeletal muscle has been investigated by means of the Calcium-stat method, in which transport may be measured in the micromolar free calcium ion concentration range, in the absence of calcium buffers. At pH 7.2 and 20 degrees C, ATP, in the range 1 to 10 mM, decreased [Ca2+]0.5 from 2.0 microM to 0.3 microM and decreased Vmax of oxalate-supported transport from 0.5 to 1.3 mumol min-1 mg-1. Simultaneous measurements of transport and of ATPase activity in the range 0.8 to 10 microM free Ca2+ showed a ratio of 2.1 calcium ions translocated/molecule of ATP hydrolyzed. Transport, in the presence of 5 mM ATP, ceased when calcium ion concentration fell to 0.6 to 1.2 microM, whilst ATPase activity of 90 nmol of ATP hydrolyzed min-1 mg-1 persisted. The data obtained by the Calcium-stat method differed from those described previously using calcium buffers, in that they showed lower apparent affinities of the transport site for calcium ions, more marked sigmoidal behavior, an effect of ATP concentration on Ca2+ concentration dependence and lower ATPase activity in the absence of transport. The calcium complex of ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (CaEGTA) had no effect when transport was stimulated maximally at saturating free Ca2+ concentrations. However, at calcium ion levels below [Ca2+]0.5, 70 microM CaEGTA stimulated transport to rates of 20 to 45% of Vmax. Half-maximal stimulation of transport occurred at 19 microM CaEGTA. CaEGTA, 50 microM, decreased [Ca2+]0.5, determined at 5 mM ATP, from 1.3 microM to 0.45 microM. It is proposed that a ternary complex, E . Ca2+ . EGTA4-, is formed as an intermediate species during CaEGTA-stimulated calcium transport by sarcoplasmic reticulum membranes and stimulates the calcium pump at limiting free Ca2+ ion concentration.     

Expression of calcium transport proteins in the extraembryonic membranes of a viviparous snake, Virginia striatula

Yolk is the primary source of calcium for embryonic growth and development for most squamates, irrespective of mode of parity. The calcified eggshell is a secondary source for embryonic calcium in all oviparous eggs, but this structure is lost in viviparous lineages. Virginia striatula is a viviparous snake in which embryos obtain calcium from both yolk and placental transport of uterine calcium secretions. The developmental pattern of embryonic calcium acquisition in V. striatula is similar to that for oviparous snakes. Calbindin-D(28K) is a marker for epithelial calcium transport activity and plasma membrane Ca(2+)-ATPase (PMCA) provides the energy to catalyze the final step in calcium transport. Expression of calbindin-D(28K) and PMCA was measured by immunoblotting in yolk sac splanchnopleure and chorioallantois of a developmental series of V. striatula to test the hypothesis that these proteins mediate calcium transport to embryos. In addition, we compared the expression of calbindin-D(28K) in extraembryonic membranes of V. striatula throughout development to a previously published expression pattern in an oviparous snake to test the hypothesis that the ontogeny of calcium transport function is independent of reproductive mode. Expression of calbindin-D(28K) increased in yolk sac splanchnopleure and chorioallantois coincident with calcium mobilization from yolk and uterine sources and with embryonic growth. The amount of PMCA in the chorioallantois did not change through development suggesting its expression is not rate limiting for calcium transport. The pattern of expression of calbindin-D(28K) and PMCA confirms our initial hypothesis that these proteins mediate embryonic calcium uptake. In addition, the developmental pattern of calbindin-D(28K) expression in V. striatula is similar to that of an oviparous snake, which suggests that calcium transport mechanisms and their regulation are independent of reproductive mode.     

Long-term prolactin exposure differentially stimulated the transcellular and solvent drag-induced calcium transport in the duodenum of ovariectomized rats

Prolactin, having been shown to stimulate transcellular active and solvent drag-induced calcium transport in the duodenum of female rats, was postulated to improve duodenal calcium transport in estrogen-deficient rats. The aim of the present study was, therefore, to demonstrate the effects of long-term prolactin exposure produced by anterior pituitary (AP) transplantation on the duodenal calcium transport in young (9-week-old) and adult (22-week-old) ovariectomized rats. We found that ovariectomy did not alter the transcellular active duodenal calcium transport in young and adult rats fed normal calcium diet (1.0% w/w Ca) but decreased the solvent drag-induced duodenal calcium transport from 75.50 +/- 10.12 to 55.75 +/- 4.77 nmol.hr(-1).cm(-2) (P < 0.05) only in adult rats. Long-term prolactin exposure stimulated the transcellular active calcium transport in young and adult AP-grafted ovariectomized rats fed with normal calcium diet by more than 2-fold from 7.56 +/- 0.79 to 16.54 +/- 2.05 (P < 0.001) and 9.78 +/- 0.72 to 15.99 +/- 1.75 (P < 0.001) nmol.hr(-1).cm(-2), respectively. However, only the solvent drag-induced duodenal calcium transport in young rats was enhanced by prolactin from 95.51 +/- 10.64 to 163.20 +/- 18.03 nmol.hr(-1).cm(-2) (P < 0.001) whereas that in adult rats still showed a decreased flux from 75.50 +/- 10.12 to 47.77 +/- 5.42 nmol.hr(-1).cm(-2) (P < 0.05). Because oral calcium supplement has been widely used to improve calcium balance in estrogen-deficient animals, the effect of a high-calcium diet (2.0% w/w Ca) was also investigated. The results showed that stimulatory action of long-term prolactin on the transcellular active duodenal calcium transport in both young and adult rats was diminished after being fed a high-calcium diet. The same diet also abolished prolactin-enhanced solvent drag-induced duodenal calcium transport in young and further decreased that in adult AP-grafted ovariectomized rats. We concluded that the solvent drag-induced duodenal calcium transport in adult rats was decreased after ovariectomy. Long-term prolactin exposure stimulated the transcellular active duodenal calcium transport in both young and adult rats whereas enhancing the solvent drag-induced duodenal calcium transport only in young rats. Effects of prolactin were abolished by a high-calcium diet.     

Calcium transport in transverse tubules isolated from rabbit skeletal muscle

Isolated transverse tubule vesicles free of sarcoplasmic reticulum transport calcium with high affinity in the presence of ATP. The calcium transport by transverse tubules differs from calcium transport by sarcoplasmic reticulum. It is not increased by oxalate or phosphate, it has a different temperature dependence, it is inhibited by sub-micromolar concentrations of orthovanadate, it is stimulated by calmodulin, and is inhibited by quercetin without causing calcium release. The rates of calcium transport by transverse tubules are two orders of magnitude lower than those of sarcoplasmic reticulum, suggesting that the calcium pump protein of transverse tubules is a minor component of the membrane. Addition of calmodulin to transverse tubule vesicles--treated with high salt in the presence of EGTA to remove endogenous calmodulin--caused a marked stimulation of transport rates at low concentrations of calcium, and decreased from 1.0 to 0.3 microM the calcium concentration at which half-maximal rates of transport were obtained. A role for the transverse tubule calcium pump in maintaining low sarcoplasmic calcium concentrations is proposed.     

Gastrointestinal calcium absorption in sheep is mostly insensitive to an alimentary induced challenge of calcium homeostasis

For ruminants, marked differences to monogastric species have been described concerning the localisation and vitamin D sensitivity of gastrointestinal calcium absorption, particularly with respect to the forestomach compartment. Therefore, we investigated gastrointestinal calcium transport of sheep as influenced by a dietary calcium restriction and/or a supraphysiological dosage of exogenous calcitriol. Using the Ussing chamber technique, we determined calcium and mannitol flux rates to differentiate between para- and transcellular calcium transport in rumen, duodenum, jejunum and colon. Expression of epithelial calcium channels, calbindin-D(9K), and basolateral extrusion mechanisms was determined by quantitative RT-PCR and Western blot analysis. Active calcium transport could be demonstrated in jejunum and rumen. A significant stimulation of jejunal calcium absorption was only observed in animals treated with calcitriol. The alimentary calcium restriction alone did not result in significant effects indicating a less effective intestinal adaptation to alimentary calcium restriction than observed in monogastric animals. The observed ruminal calcium transport was not affected at all, neither by the diet nor the calcitriol treatment. Furthermore, no significant expression of epithelial calcium channels or calbindin-D(9K) could be detected in the rumen; therefore it is concluded that calcium transport in the forestomachs is probably mediated by a different, so far unknown mechanism.     

Sources and timing of calcium mobilization during embryonic development of the corn snake, Pantherophis guttatus

Embryos of oviparous Reptilia (=turtles, lepidosaurs, crocodilians and birds) extract calcium for growth and development from reserves in the yolk and eggshell. Yolk provides most of the calcium to embryos of lizards and snakes. In contrast, the eggshell supplies most of the calcium for embryonic development of turtles, crocodilians and birds. The yolk sac and chorioallantoic membrane of birds recover and transport calcium from the yolk and eggshell and homologous membranes of squamates (lizards and snakes) probably transport calcium from these two sources as well. We studied calcium mobilization by embryos of the snake Pantherophis guttatus during the interval of greatest embryonic growth and found that the pattern of calcium transfer was similar to other snakes. Calcium recovery from the yolk is relatively low until the penultimate embryonic stage. Calcium removal from the eggshell begins during the same embryonic stage and total eggshell calcium drops in each of the final 2 weeks prior to hatching. The eggshell supplies 28% of the calcium of hatchlings. The timing of calcium transport from the yolk and eggshell is coincident with the timing of growth of the yolk sac and chorioallantoic membrane and expression of the calcium binding protein, calbindin-D28K, in these tissues as reported in previous studies. In the context of earlier work, our findings suggest that the timing and mechanism of calcium transport from the yolk sac of P. guttatus is similar to birds, but that both the timing and mechanism of calcium transport by the chorioallantoic membrane differs. Based on the coincident timing of eggshell calcium loss and embryonic calcium accumulation, we also conclude that recovery of eggshell calcium in P. guttatus is regulated by the embryo.     

Compartmentation of calcium in digitonin-disrupted guinea pig pancreatic acinar cells

The treatment of guinea pig pancreatic acinar cells with digitonin leads to disruption of the plasma membrane, as judged by the liberation of cytosolic enzymes, without significant alteration of the mitochondrial membrane. The transport of calcium by the particulate residue was studied, and two different pools could be distinguished. One was supported by ATP or ADP, succinate providing the respiratory substrate, and was sensitive to the inhibitors, Ruthenium red and azide. The other pool needed the presence of ATP, ADP being ineffective, and also was unaffected by Ruthenium red or by azide, but was stimulated several-fold by oxalate. The Ruthenium red-sensitive calcium pool has characteristics resembling those of the transport of calcium by a mitochondrial fraction prepared from digitonin-treated acinar cells. In contrast, the Ruthenium red-insensitive calcium transport has characteristics resembling those of a microsomal fraction obtained from guinea pig pancreas. When the transport of calcium in digitonized cells was assayed at a calcium concentration range of 10(-8)-10(-4) M, preferential Ruthenium red-insensitive calcium transport could be observed at submicromolar calcium concentrations.     

A role for calcium in auxin transport

The basipetal transport of the auxin, indoleacetic acid, in sunflower stem sections is markedly suppressed by washing the tissue in ethylenediaminetetraacetate, and transport is restored by subsequent application of calcium solutions. The ethylenediaminetetraacetate treatment is shown to result in the removal of substantial amounts of calcium from the tissue, and the restoration of transport is distinctive for calcium solutions, lesser effects being observed for magnesium and lanthanum, and little effect for monovalent cations. The calcium effects are interpreted as indicating that the auxin transport system depends upon structural or functional features of cellular membranes which involve calcium in a manner analogous to the transport of inorganic ions.     

Regulation of cardiac sarcoplasmic reticulum calcium transport by calcium-calmodulin-dependent phosphorylation

Cardiac sarcoplasmic reticulum contains an endogenous calcium-calmodulin-dependent protein kinase and a 22,000-Da substrate, phospholamban. This kinase is half-maximally activated (EC50) by 3.8 +/- 0.3 microM calcium and is absolutely dependent on exogenous calmodulin (EC50 = 49 nM). To determine the effect of this phosphorylation on calcium transport, sarcoplasmic reticulum vesicles (0.5 mg/ml) were preincubated under conditions for optimal phosphorylation (50 mM potassium phosphate, pH 7.0, 10 mM MgCl2, 0.5 mM EGTA, 0.478 mM CACl2, 0.1 microM calmodulin, 0.5 mM ATP). Control sarcoplasmic reticulum was preincubated under identical conditions but in the absence of ATP to avoid phosphorylation. Both control and phosphorylated vesicles were centrifuged and resuspended in 0.3 M sucrose, 20 mM Tris-HCl, 100 mM KCl, pH 7.0, to remove calmodulin and subsequently assayed for calcium (45Ca) transport in the presence of 2.5 mM Tris-oxalate. Phosphorylation of sarcoplasmic reticulum vesicles by calcium-calmodulin-dependent protein kinase resulted in a significant increase (2- to 4-fold) in the rate of calcium transport at low calcium concentrations (less than 3 microM), while calcium transport was minimally affected at higher calcium. Hill coefficients (n) derived from Hill plots of transport data showed no difference between control and phosphorylated sarcoplasmic reticulum (n = 2.0), indicating that phosphorylation does not alter the cooperativity between calcium sites on the calcium pump. The EC50 for calcium activation of calcium transport by control vesicles was 0.86 +/- 0.1 microM calcium, and phosphorylation of phospholamban decreased this value to 0.61 +/- 0.07 microM calcium (n = 7, p less than 0.028), indicating an increase in the apparent affinity for calcium upon phosphorylation. These results were found to be specific for calcium-calmodulin-dependent phosphorylation of phospholamban. Control experiments on the effects of the reactants used in the phosphorylation assay and subsequent centrifugation of sarcoplasmic reticulum showed no alteration of the rate of calcium transport. Therefore, the calcium pump in cardiac sarcoplasmic reticulum appears to be regulated by an endogenous calcium-calmodulin-dependent protein kinase, and this may provide an important regulatory mechanism for the myocardium.     

Expression of calbindin-D28K by yolk sac and chorioallantoic membranes of the corn snake, Elaphe guttata

The yolk splanchnopleure and chorioallantoic membrane of oviparous reptiles transport calcium from the yolk and eggshell to the developing embryo. Among oviparous amniotes, the mechanism of calcium mobilization to embryos has been studied only in domestic fowl, in which the mechanism of calcium transport of the yolk splanchnopleure differs from the chorioallantoic membrane. Transport of calcium is facilitated by calbindin-D(28K) in endodermal cells of the yolk splanchnopleure of chickens but the chorioallantoic membrane does not express calbindin-D(28K). We used immunoblotting to assay for calbindin-D(28K) expression in yolk splanchnopleure and chorioallantoic membrane of the corn snake, Elaphe guttata, to test the hypothesis that the mechanism of calcium transport by extraembryonic membranes of snakes is similar to birds. High calbindin-D(28K) expression was detected in samples of yolk splanchnopleure and chorioallantoic membrane during late embryonic stages. We conclude that calbindin-D(28K) is expressed in these extraembryonic membranes to facilitate transport of calcium and that the mechanism of calcium transport of the chorioallantoic membrane of the corn snake differs from that of the chicken. Further, we conclude that calbindin-D(28K) expression is developmentally regulated and increases during later embryonic stages in the corn snake.     

Glucose 1-phosphate increases active transport of calcium in intestine

Active calcium transport in intestine is essential for serum calcium homeostasis as well as for bone formation. It is well recognized that vitamin D is a major, if not sole, stimulator of intestinal calcium transport activity in mammals. Besides vitamin D, endogenous glucose 1-phosphate (G1P) affects calcium transport activity in some microorganisms. In this study, we investigated whether G1P affects intestinal calcium transport activity in mammals as well. Of several glycolytic intermediates, G1P was the sole sugar compound in stimulating intestinal calcium uptake in Caco-2 cells. G1P stimulated net calcium influx and expression of calbindin D9K protein in rat intestine, through an active transport mechanism. Calcium uptake in G1P-supplemented rats was greater than that in the control rats fed a diet containing adequate vitamin D3. Bone mineral density (BMD) of aged rat femoral metaphysis and diaphysis was also increased by feeding the G1P diet. G1P did not affect serum levels of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] at all. These results suggest that exogenously applied G1P stimulates active transport of calcium in intestine, independent of vitamin D, leading to an increase of BMD.     

Routes of calcium movement across the chick chorioallantois.

The mechanism of calcium transport across the chick chorioallantois has been studied in vivo. The 15 day old membrane contains no calcium binding protein (CaBP) and attempts to stimulate transport by adding this protein or vitamin D metabolites did not enhance the normal transport process. The mitochondrial and microsomal fractions of transporting cells contain only a small percentage of the 45Ca actually moved across the epithelium and simultaneous measurements of 45Ca and 3H inulin indicate that the bulk movement of fluid is not involved in calcium transport. Calcium appears to be moved after being bound to a protein since strontium and PCMBS both interfere with transport and the membrane shows saturation kinetics. It is suggested that none of the existing theories provides an adequate explanation for the mechanism of calcium transport and some support is given to the possibility that intercellular routes are involved in part of the process.     

Comparison of the properties of active Ca2+ transport by the islet-cell endoplasmic reticulum and plasma membrane

The properties of active or ATP-dependent calcium transport by islet-cell endoplasmic reticulum and plasma membrane-enriched subcellular fractions were directly compared. These studies indicate that the active calcium transport systems of the two membranes are fundamentally distinct. In contrast to calcium uptake by the endoplasmic reticulum-enriched fraction, calcium uptake by islet-cell plasma membrane-enriched vesicles exhibited a different pH optimum, was not sustained by oxalate, and showed an approximate 30-fold greater affinity for ionized calcium. A similar difference in affinity for calcium was exhibited by the Ca²⁺-stimulated ATPase activities which are associated with these islet-cell subcellular fractions. Consistent with the effects of calmodulin on calcium transport, calmodulin stimulated Ca²⁺-ATPase in the plasma membranes, but did not increase calcium-stimulated ATPase activity in the endoplasmic reticulum membranes. The physiological significance of the differences observed in calcium transport by the endoplasmic reticulum and plasma membrane fractions relative to the regulation of insulin secretion by the islets of Langerhans is discussed.     

Functional analysis of placental 57-kDa Ca(2+)-binding protein: overexpression and downregulation in a trophoblastic cell line.

The placental trophoblastic epithelium functions to transport nutrients needed by the fetus, including calcium, which is required in the greatest amounts during the last third of pregnancy when the majority of fetal skeletal mineralization occurs. The mechanism of placental calcium transport and the developmental changes in the trophoblast that facilitate this process are currently incompletely understood. We have previously identified a 57-kDa, Ca(2+)-binding protein (CaBP) functionally implicated in placental calcium transport and trophoblast differentiation. In this study we have directly examined the role of CaBP in these processes by (1) recombinantly overexpressing CaBP in an inducible manner and (2) downregulating CaBP expression using antisense technology, using the rat choriocarcinoma cell line Rcho-1 as a trophoblastic cell model system. Our results show that overexpression of CaBP stimulates both cellular calcium uptake and vectorial calcium transport activities in Rcho-1 cells. Those cells stably expressing CaBP also exhibit higher levels of steady-state intracellular calcium and enhanced calcium-buffering ability. In addition, prolonged overexpression of CaBP in Rcho-1 cultures promotes trophoblast differentiation. Conversely, downregulation of CaBP expression had a negative effect on calcium uptake, calcium transport, and trophoblast differentiation in Rcho-1 cells. These data indicate that CaBP plays a direct role in placental calcium transport, functioning both as an intracellular calcium buffer and as a shuttle. These results also support a more direct role for CaBP in the trophoblast differentiation pathway.     

A role of calcium in altered sodium ion transport of hypertensives?

Research on the etiology of essential hypertension has led to many reports of altered ion transport in cells from hypertensive patients and animal models. Abnormalities in sodium and calcium ion gradients and transport in vascular smooth muscle, neuronal tissue, cardiac muscle as well as erythrocytes have been extensively investigated. It is not clear whether these abnormalities are of primary or secondary nature. The current knowledge of sodium and calcium ion transport in essential hypertension is briefly reviewed here. Furthermore, evidence is presented which suggests a role of calcium in the regulation of sodium transport activity.