Net Fluxes Of Ca2 Research Articles

Transcriptome analysis of Gossypium reveals the molecular mechanisms of Ca2+ signaling pathway on arsenic tolerance induced by arbuscular mycorrhizal fungi.

Arbuscular mycorrhizal fungi (AMF) have been demonstrated their ability to enhance the arsenic (As) tolerance of host plants, and making the utilization of mycorrhizal plants a promising and practical approach for remediating As-contaminated soils. However, comprehensive transcriptome analysis to reveal the molecular mechanism of As tolerance in the symbiotic process between AMF and host plants is still limited. In this study, transcriptomic analysis of Gossypium seedlings was conducted with four treatments: non-inoculated Gossypium under non-As stress (CK0), non-inoculated Gossypium under As stress (CK100), F. mosseae-inoculated Gossypium under non-As stress (FM0), and F. mosseae-inoculated Gossypium under As stress (FM100). Our results showed that inoculation with F. mosseae led to a reduction in net fluxes of Ca2+, while increasing Ca2+ contents in the roots and leaves of Gossypium under the same As level in soil. Notably, 199 and 3129 differentially expressed genes (DEGs) were specially regulated by F. mosseae inoculation under As stress and non-As stress, respectively. Through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation and enrichment analyses, we found that under As stress, F. mosseae inoculation up-regulated a significant number of genes related to the Ca2+ signaling pathway genes, involved in cellular process, membrane part, and signal transduction. This suggests a potential role in mitigating As tolerance in Gossypium seedlings. Furthermore, our analysis identified specific DEGs in transcription factor families, including ERF, MYB, NAC, and WRKY, that were upregulated by F. mosseae inoculation. Conversely, MYB and HB-other were down-regulated. The ERF and MYB families exhibited the highest number of up- and down-regulated DEGs, respectively, which were speculated to play an important role in alleviating the As toxicity of Gossypium. Our findings provided valuable insights into the molecular theoretical basis of the Ca2+ signaling pathway in improving As tolerance of mycorrhizal plants in the future.

Physiological and molecular mechanisms mediating xylem Na+ loading in barley in the context of salinity stress tolerance

Time-dependent kinetics of xylem Na+ loading was investigated using a large number of barley genotypes contrasting in their salinity tolerance. Salt-sensitive varieties were less efficient in controlling xylem Na+ loading and showed a gradual increase in the xylem Na+ content over the time. To understand underlying ionic and molecular mechanisms, net fluxes of Ca2+ , K+ and Na+ were measured from the xylem parenchyma tissue in response to H2 O2 and ABA; both of them associated with salinity stress signalling. Our results indicate that NADPH oxidase-mediated apoplastic H2 O2 production acts upstream of the xylem Na+ loading and is causally related to ROS-inducible Ca2+ uptake systems in the root stelar tissue. It was also found that ABA regulates (directly or indirectly) the process of Na+ retrieval from the xylem and the significant reduction of Na+ and K+ fluxes induced by bumetanide are indicative of a major role of chloride cation co-transporter (CCC) on xylem ion loading. Transcript levels of HvHKT1;5_like and HvSOS1_like genes in the root stele were observed to decrease after salt stress, while there was an increase in HvSKOR_like gene, indicating that these ion transporters are involved in primary Na+ /K+ movement into/out of xylem.

Nitrate supply affects ammonium transport in canola roots

Plants may suffer from ammonium (NH4+) toxicity when NH4+ is the sole nitrogen source. Nitrate (NO3-) is known to alleviate NH4+ toxicity, but the mechanisms are unknown. This study has evaluated possible mechanisms of NO3- alleviation of NH4+ toxicity in canola (Brassica napus L.). Dynamics of net fluxes of NH4+, H+, K+ and Ca2+ were assessed, using a non-invasive microelectrode (MIFE) technique, in plants having different NO3- supplies, after single or several subsequent increases in external NH4Cl concentration. After an increase in external NH4Cl without NO3-, NH4+ net fluxes demonstrated three distinct stages: release (tau1), return to uptake (tau2), and a decrease in uptake rate (tau3). The presence of NO3- in the bathing medium prevented the tau1 release and also resulted in slower activation of the tau3 stage. Net fluxes of Ca2+ were in the opposite direction to NH4+ net fluxes, regardless of NO3- supply. In contrast, H+ and K+ net fluxes and change in external pH were not correlated with NH4+ net fluxes. It is concluded that (i) NO3- primarily affects the NH4+ low-affinity influx system; and (ii) NH4+ transport is inversely linked to Ca2+ net flux.

Osmotic sensitivity of Ca2+ and H+ transporters in corn roots: effect on fluxes and their oscillations in the elongation region.

Seedling roots of corn were treated with different concentrations of mannitol-containing solution for 1 to 1.5 hr, and net fluxes of Ca2+ and H+ were measured in the elongation region. H+ fluxes were much more sensitive to osmotic pressure than were Ca2+ fluxes. Oscillations of 7-min period in H+ flux, normally observed in the control, were almost fully suppressed at high osmotic concentrations. Net H+ flux was shifted from average efflux of 25 +/- 3 nmol m-2 sec-1 to average influx of 10 +/- 5 nmol m-2 sec-1 after the incubation in 100 mm mannitol. The larger the osmotic concentration, the larger was the H+ influx. This flux caused the unbuffered solution of pH 4.85 to change to pH 5.3 after mannitol application. It appears that the osmoticum suppresses oscillatory H+ extrusion at the plasma membrane. Discrete Fourier Transforms of the H+ flux data showed that, apart from suppression of the 7-min oscillations in H+ flux, mannitol also promoted the appearance of faster 2-min oscillations. Ca2+ influx slightly increased after mannitol treatment. In addition the 7-min oscillatory component of Ca2+ flux remained apparent thereby showing independence of H+ flux.

Calcium transport across the isolated oral epithelium of scleractinian corals

Oral epithelia were isolated from Lobophyllia temprichii and Plerogyra sinuosa and placed in Ussing chambers. Calcium flux was measured under open circuit and short circuit conditions using 45Ca. Only a small transepithelial potential of 1.5 mV was recorded under open circuit conditions and no effect on flux rates were observed when the preparation was short circuited. Unidirectional fluxes in single and paired experiments were consistently greater in the ectoderm to gastroderm direction than from gastroderm to ectoderm with net flux of Ca2+ frequently being more than 3x10-4 μEq mm-2 min-1. A small number of paired experiments showed that net flux of Ca2+ was reduced by Sr2+ and sodium azide but not by dinitrophenol. Unidirectional fluxes from ectoderm to gastroderm appeared to have maxima in the early and late parts of the day when recorded between 0900 and 2100 hrs. It is concluded that active transport of Ca2+ occurs across the isolated oral epitheia and that this may be an initial step in the process of keletal Ca2+ deposition.

Flux of calcium across the sheep rumen wall in vivo and in vitro.

Experiments were carried out in vivo and in vitro to study the flux of Ca2+ across the sheep rumen wall. Buffer solutions with five different Ca2+ concentrations in the range 0.2-3.6 mmol l-1 were introduced into the isolated, emptied and washed reticulo-rumen for 1 h to determine Ca2+ net flux. Phosphate concentrations were either kept constant at 8 mmol l-1, or varied so as to maintain a constant phosphate/Ca2+ ratio of 2:1. Under both conditions there was a Ca2+ net secretion into the rumen at Ca2+ concentrations below approximately 1 mmol l-1 whereas Ca2+ net absorption occurred at higher Ca2+ concentrations and linearly increased with increasing Ca2+ concentrations. When phosphate concentrations were reduced to 4.3 and 0.6 mmol l-1 (Ca2+, 3 mmol l-1) Ca2+ net absorption was reduced by about 50% and completely abolished, respectively. The unidirectional (mucosal-to-serosal or serosal-to-mucosal) Ca fluxes in vitro, JCams and JCasm, were measured in Ussing chambers with computer-controlled voltage clamp. Under short-circuit current conditions (Ca2+ and phosphate, 2 mmol l-1) there was a significant net flux of Ca2+ from the mucosal to the serosal side. This net flux of Ca2+ was abolished when ouabain was added to the serosal solution, indicating that the unidirectional Ca2+ flux JCams and, hence, the net flux of Ca2+ was dependent upon active Na+ transport accomplished by the Na, K-ATPase system. Under voltage clamp conditions, the net flux of Ca2+ was greatly increased at a potential difference PD = -25 mV (serosa negative) but disappeared at PD = +25 mV (serosa positive). Analysis of the unidirectional Ca2+ fluxes JCams and JCasm under varying potential differences indicated that both Ca2+ fluxes consisted of a PD-dependent and a PD-independent component. The results show that the reticulo-rumen participates in Ca2+ absorption in sheep.

Glucose-stimulated sequestration of Ca2+ in clonal insulin-releasing cells. Evidence for an opposing effect of muscarinic-receptor activation.

Net fluxes of Ca2+ and acid production were studied in clonal insulin-releasing cells (RINm5F) by using colour indicators and dual-wavelength spectrophotometry. After equilibration with a medium containing 10-20 microM-Ca2+, only minimal amounts of Ca2+ (0.08 mmol/kg of protein) were released from the cells by subsequent additions of the respiratory blocker antimycin A and the Ca2+ ionophore A23187. The presence of 20 mM-glucose resulted in an almost 5-fold increase of the acid production and in a stimulated net uptake of Ca2+. The latter process was independent of the extracellular Ca2+ concentration and reached saturation after 20 +/- 1 min, when it corresponded to 1.18 +/- 0.07 mmol of calcium/kg of protein. Whereas the thiol reagent iodoacetamide suppressed the acid production, interference with mitochondrial function by using antimycin A or the uncoupler carbonyl cyanide m-chlorophenylhydrazone had the opposite effect. The latter two drugs induced a selective release of Ca2+ from a pool containing 35% of that taken up during glucose exposure. Most of the remaining Ca2+ was liberated by A23187 or iodoacetamide. Carbamoylcholine was also selective in mobilizing glucose-stimulated calcium, but this calcium (17%) appeared to originate from the pool insensitive to mitochondrial poisons. The action of carbamoylcholine was blocked by atropine and did not depend on the presence of extracellular Na+. The opposite effects of glucose and muscarinic-receptor activation on a non-mitochondrial calcium pool are consistent with participation of the endoplasmic reticulum in the glucose-induced sequestration of Ca2+ in pancreatic beta-cells.

Inhibition of net calcium efflux from bone by ethanol in vitro.

Ethanol administered to animals is known to cause hypocalcemia. This investigation was undertaken to evaluate direct early effects of ethanol on the net fluxes of Ca2+ and inorganic phosphate (Pi) into or out of bone using tibiae from 13-day-old chick embryos and 8-day-old suckling rats in vitro. When chick bones were incubated with ethanol in the medium (10 microliters/ml), net Ca2+ efflux was decreased 19, 22, 27, and 31% at 2, 4, 6, and 8 h, respectively. Metabolically inhibited bones showed a net influx rather than efflux of Ca2+ and were not further affected by ethanol. The ethanol had no consistent effect on net Pi efflux and slightly reduced lactate production. At three doses of ethanol (3, 10, and 30 microliters/ml of medium) the reduction of Ca2+ efflux was dose related in both chick and rat bones. Ethanol completely inhibited parathyroid hormone-stimulated, net Ca2+ efflux from chick bones, even at ethanol levels that by themselves did not affect Ca2+ efflux. These findings are consistent with the hypothesis that the hypocalcemia in ethanol-treated animals may be due in part to inhibition of net Ca2+ efflux from bone.

Depolarization-independent net uptake of calcium into clonal insulin-releasing cells exposed to glucose.

Insulin release, net fluxes of Ca2+, and glucose metabolism were studied in a clonal cell line (RINm5F) established from a transplantable rat islet tumor. The insulin content amounted to only 0.03% of that of the total protein and decreased even further with subsequent passages. The insulin secretion was as high as 10 to 20% of the total hormone content per hour. Insulin release was stimulated by K+ depolarization but not by exposure to glucose. In contrast to this secretory pattern, glucose but not K+ stimulated the net uptake of Ca2+ at micromolar concentrations of the ion. The glucose effect was not mimicked by 20 mM 3-O-methylglucose. It was as pronounced at 1 mM as at 20 mM of the sugar and corresponded to an uptake of 119 fmol cm-2 s-1. Glucose metabolism was typical for tumor cells with a high glycolytic flux and an oxidation-to-utilization ratio as low as 0.05-0.15. Maximal oxidative degradation was attained already at 1 mM. This concentration was also equivalent to the Km for glucose utilization, indicating a substantial left-hand shift of the normal dose-response curve. It is suggested that glucose induces a depolarization-independent net uptake of Ca2+ by favouring intracellular buffering of the cation.

Evidence for more than one Ca2+ transport mechanism in mitochondria.

The active transport and internal binding of the Ca2+ analogue Mn2+ by rat liver mitochondria were monitored with electron paramagnetic resonance. The binding of transported Mn2+ depended strongly on internal pH over the range 7.7-8.9. Gradients of free Mn2+ were compared with K+ gradients measured on valinomycin-treated samples. In the steady state, the electrochemical Mn2+ activity was larger outside than inside the mitochondria. The observed gradients of free Mn2+ and of H+ could not be explained by a single "passive" uniport or antiport mechanism of divalent cation transport. This conclusion was further substantiated by observed changes in steady-state Ca2+ and Mn2+ distributions induced by La3+ and ruthenium red. Ruthenium red reduced total Ca2+ or Mn2+ uptake, and both inhibitors caused release of divalent cation from preloaded mitochondria. A model is proposed in which divalent cations are transported by at least two mechanisms: (1) a passive uniport and (2) and active pump, cation antiport or anion symport. The former is more sensitive to La3+ and ruthenium red. Under energized steady-state conditions, the net flux of Ca2+ or Mn2+ is inward over (1) and outward over (2). The need for more than one transport system inregulating cytoplasmic Ca2+ is discussed.