Searching for a Role of NCX/NCKX Exchangers in Neurodegeneration

Control of intracellular calcium signaling is essential for neuronal development and function. Maintenance of Ca²⁺ homeostasis depends on the functioning of specific transport systems that remove calcium from the cytosol. Na⁺/Ca²⁺ exchange is the main calcium export mechanism across the plasma membrane that restores resting levels of calcium in neurons after stimulation. Two families of Na⁺/Ca²⁺ exchangers exist, one of which requires the co-transport of K⁺ and Ca²⁺ in exchange for Na⁺ ions. The malfunctioning of Na⁺/Ca²⁺ exchangers has been related to the development of pathological conditions in the regulation of neuronal death after hypoxia–anoxia, brain trauma, and nerve injury. In addition, the Na⁺/Ca²⁺ exchanger function has been associated with impaired Ca²⁺ homeostasis during aging of the brain, as well as with a role in Alzheimer’s disease by regulating β-amyloid toxicity. In this review, we summarize the current knowledge about the Na⁺/Ca²⁺ exchanger families and their implications in neurodegenerative disorders.     

Kinetic properties of Na+/Ca2+ exchange in basolateral plasma membranes of rat small intestine

The presence of an Na⁺/Ca²⁺ exchange system in basolateral plasma membranes from rat small intestinal epithelium has been demonstrated by studying Na⁺ gradient-dependent Ca²⁺ uptake and the inhibition of ATP-dependent Ca²⁺ accumulation by Na⁺. The presence of 75 mM Na⁺ in the uptake solution reduces ATP-dependent Ca²⁺ transport by 45%, despite the fact that Na⁺ does not affect Ca²⁺-ATPase activity. Preincubation of the membrane vesicles with ouabain or monensin reduces the Na⁺ inhibition of ATP-dependent Ca²⁺ uptake to 20%, apparently by preventing accumulation of Na⁺ in the vesicles realized by the Na⁺-pump. It was concluded that high intravesicular Na⁺ competes with Ca²⁺ for intravesicular Ca²⁺ binding sites. In the presence of ouabain, the inhibition of ATP-dependent Ca²⁺ transport shows a sigmoidal dependence on the Na⁺ concentration, suggesting cooperative interaction between counter transport of at least two sodium ions for one calcium ion. The apparent affinity for Na⁺ is between 15 and 20 mM. Uptake of Ca²⁺ in the absence of ATP can be enhanced by an Na⁺ gradient (Na⁺ inside > Na⁺ outside). This Na⁺ gradient-dependent Ca²⁺ uptake is further stimulated by an inside positive membrane potential but abolished by monensin. The apparent affinity for Ca²⁺ of this system is below 1 μM. In contrast to the ATP-dependent Ca²⁺ transport, there is no significant difference in Na⁺ gradient-dependent Ca²⁺ uptake between basolateral vesicles from duodenum, midjejunum and terminal ileum. In duodenum the activity of ATP-driven Ca²⁺ uptake is 5-times greater than the Na⁺/Ca²⁺ exchange capacity but in the ileum both systems are of equal potency. Furthermore, the Na⁺/Ca²⁺ exchange mechanism is not subject to regulation by 1α,25-dihydroxy vitamin D-3, since repletion of vitamin D-deficient rats with this seco-steroid hormone does not influence the Na⁺/Ca²⁺ exchange system while it doubles the ATP-driven Ca²⁺ pump activity.     

Incorporation of the transmembrane hydrophobic domain of glycophorin into small unilamellar phospholipid vesicles Ion Flux studies

Human erythrocyte glycophorin is one of the best characterized integral membrane proteins. Reconstitution of the membrane-spanning hydrophobic segment of glycophorin (the tryptic insoluble peptide released when glycophorin is treated with trypsin) with liposomes results in the production of freeze-fracture intrabilayer particles of 80 Å diameter (Segrest, J.P., Gulik-Krzywicki, T. and Sardet, C. (1974) Proc. Natl. Acad. Sci. U.S.A. 71, 3294–3298), with particles appearing at or above a tryptic insoluble peptide concentration of 4 mmol per mol phosphatidylcholine. In the present study, increasing concentrations of tryptic insoluble peptide were added to sonicated small unilamellar egg phosphatidylcholine vesicles and the rate of efflux of ²²Na⁺ was examined by rapid (30 s) gel filtration on Sephadex G-50. Below a concentation of 3–5 mmol tryptic insoluble peptide/mol phosphatidylcholine, ²²Na⁺ efflux occurs at a constant slow rate at given tryptic insoluble peptide concentrations. Above a concentration of 3–5 mM, the rate of efflux is biphasic at given tryptic insoluble peptide concentrations, exhibiting both an initial fast and a subsequent slow component. On the basis of graphic and computer curve-fitting analysis, with increasing tryptic insoluble peptide concentration, the rate of the slow component reaches a plateau at a tryptic insoluble peptide concentration of 3–5 mM and remains essentially constant until much higher concentrations are reached; the fast component increases linearly with increasing tryptic insoluble peptide concentration well beyond 5 mM. The most consistent interpretation of this data is as follows. The slow ²²Na⁺ efflux component is due to perturbations of small unilamellar vesicle integrity by tryptic insoluble peptide monomers. At a tryptic insoluble peptide concentration of 3–5 mmol/mol, a critical concentration is reached following which there is intrabilayer tryptic insoluble peptide self-association. The fast ²²Na⁺ efflux component is due to the increasing presence of tryptic insoluble peptide self-associated multimers the 80-Å particles seen by freeze-fracture electron microscopy) which results in a significantly larger bilayer defect than do tryptic insoluble peptide monomers. The failure of complete saturation of efflux by the fast component is ascribed to the presence of two populations of small unilamellar vesicles, some of which contain tryptic insoluble peptide multimers and some of which do not.Addition of cholesterol to the tryptic insoluble peptide/phosphatidylcholine vesicles decreases the rate of ²²Na⁺ efflux by inhibiting primarily the fast component. Freeze-fracture electron microscopy indicates that the presence of cholesterol has no effect on the size, number or distribution of 80-Å intra-bilayer particles in the tryptic insoluble peptide/phosphatidylcholine vesicles. These results are consistent with a mechanism to explain the fast Na⁺ efflux component involving protein-lipid boundary perturbations.Efflux of ⁴⁵Ca²⁺ from phosphatidylcholine vesicles is also enhanced by incorporation of tryptic insoluble peptide, but only if divalent cations (Ca²⁺ or Mg²⁺) are present in the external bathing media as well as inside the sonicated vesicles. If monovalent Na⁺ only is present in the bathing media no ⁴⁵Ca²⁺ efflux is seen. Under conditions where ⁴⁵Ca²⁺ efflux is seen, both a fast and a slow component are present, although both appear lower than corresponding rate constants for ²²Na⁺ efflux. These results suggest a coordinated mechanism for ion efflux induced by tryptic insoluble peptide and, together with the ²²Na⁺ efflux studies, may have mechanistic implications for the transbilayer phospholipid exchange (flip-flop) suggesed to be induced at glycophorin/phospholipid interfaces (de Kruiff, B., van Zoelen, E.J.J. and van Deenen, L.L.M. (1978) Biochim. Biophys. Acta 509, 537–542).     

Modulation of elastase-like activity in fibroblasts stimulated with elastin peptides

Elastin-derived peptides, kappa-elastin, prepared by chemical degradation of insoluble elastin from bovine ligamentum nuchae, were shown to increase the elastase-like activity in the culture medium and cell fractions in fibroblasts. Preincubation of cells with nifedipine (calcium channel blocker) and trifluoperazine (calmodulin antagonist) induced a decrease in the activities of the enzyme under study. These data suggest the possibility of pharmacological modulation of the biological effects induced by elastin-derived peptides.     

Seeing the Forest through the Trees: towards a Unified View on Physiological Calcium Regulation of Voltage-Gated Sodium Channels

Voltage-gated sodium channels (Na_Vs) underlie the upstroke of the action potential in the excitable tissues of nerve and muscle. After opening, Na_Vs rapidly undergo inactivation, a crucial process through which sodium conductance is negatively regulated. Disruption of inactivation by inherited mutations is an established cause of lethal cardiac arrhythmia, epilepsy, or painful syndromes. Intracellular calcium ions (Ca²⁺) modulate sodium channel inactivation, and multiple players have been suggested in this process, including the cytoplasmic Na_V C-terminal region including two EF-hands and an IQ motif, the Na_V domain III-IV linker, and calmodulin. Calmodulin can bind to the IQ domain in both Ca²⁺-bound and Ca²⁺-free conditions, but only to the DIII-IV linker in a Ca²⁺-loaded state. The mechanism of Ca²⁺ regulation, and its composite effect(s) on channel gating, has been shrouded in much controversy owing to numerous apparent experimental inconsistencies. Herein, we attempt to summarize these disparate data and propose a novel, to our knowledge, physiological mechanism whereby calcium ions promote sodium current facilitation due to Ca²⁺ memory at high-action-potential frequencies where Ca²⁺ levels may accumulate. The available data suggest that this phenomenon may be disrupted in diseases where cytoplasmic calcium ion levels are chronically high and where targeted phosphorylation may decouple the Ca²⁺ regulatory machinery. Many Na_V disease mutations associated with electrical dysfunction are located in the Ca²⁺-sensing machinery and misregulation of Ca²⁺-dependent channel modulation is likely to contribute to disease phenotypes.     

Ionic regulation of the cardiac sodium-calcium exchanger

The Na⁺-Ca²⁺ exchanger (NCX) links transmembrane movements of Ca²⁺ ions to the reciprocal movement of Na+ ions. It normally functions primarily as a Ca²⁺ efflux mechanism in excitable tissues such as the heart, but it can also mediate Ca²⁺ influx under certain conditions. Na⁺ and Ca²⁺ ions exert complex regulatory effects on NCX activity. Ca²⁺ binds to two regulatory sites in the exchanger's central hydrophilic domain, and this interaction is normally essential for activation of exchange activity. High cytosolic Na⁺ concentrations, however, can induce a constitutive activity that by-passes the need for allosteric Ca²⁺ activation. Constitutive NCX activity can also be induced by high levels of phopshotidylinositol-4,5-bisphosphate (PIP₂) and by mutations affecting the regulatory calcium binding domains. In addition to promoting constitutive activity, high cytosolic Na⁺ concentrations also induce an inactivated state of the exchanger (Na⁺-dependent inactivation) that becomes dominant when cytosolic pH and PIP₂ levels fall. Na+-dependent inactivation may provide a means of protecting cells from Ca²⁺ overload due to NCX-mediated Ca²⁺ influx during ischemia.     

Measurements of intracellular ionized calcium in squid giant axons using calcium-selective electrodes

Ca²⁺-selective electrodes have been used to measure free intracellular Ca²⁺ concentrations in squid giant axons. Electrodes made of glass cannulas of about 20 μm in diameter, plugged with a poly(vinyl chloride) gelled sensor were used to impale the axons axially. They showed a Nernstian response to Ca²⁺ down to about 3 μM in solutions containing 0.3 M K⁺ and 0.025 M Na⁺. Sub-Nernstian but useful responses were obtained up to pCa 8. The electrodes showed adequate selectivity to Ca²⁺ over Mg²⁺, H⁺, K⁺ and Na⁺. To calibrate them properly, a set of standard solutions were prepared using different Ca²⁺ buffers (EGTA, HEEDTA, nitrilotriacetic acid) after carefully characterizing their apparent Ca²⁺ association constants under conditions resembling the axoplasmic environment. In fresh axons incubated in artificial seawater containing 4 mM Ca²⁺, the mean resting intracellular ionized calcium concentration was 0.106 μM ($\text{n = 15}$). The Ca²⁺-electrodes were used to investigate effects of different experimental procedures on the [Ca²⁺]_i. The main conclusions are: (i) intact axons can extrude calcium ions at low [Ca²⁺]_i levels by a process independent of external Na⁺; (ii) poisoned axons can extrude calcium ions at high levels of [Ca²⁺]_i by an external Na⁺-dependent process. The level of free intracellular Ca attained at these latter conditions is about an order to magnitude greater than the resting physiological value.     

An investigation on calcium taurodeoxycholate micellar aggregates

Abstract

The formation of micellar aggregates in the presence of calcium(II) ions in solutions containing sodium and taurodeoxycholate ions and their composition at 25°C and in 0.5 mol dm^?3 N(CH₃)₄Cl as constant ionic medium was studied. The study was carried out by means of two different procedures. In the first one, solid calcium oxalate was equilibrated with taurodeoxycholate, sodium and hydrogen ions and the free concentration of sodium and hydrogen ions was determined. After filtration, the calcium(II) (by atomic absorption spectrophotometry) and oxalate concentration were also determined. In the second approach, hydrogen and sodium ions free concentrations were obtained by electromotive force measurements carried out in solutions containing taurodeoxycholate. The results of both procedures could be explained by assuming the presence of aggregates of different composition with the participation of sodium, calcium(II) and taurodeoxycholate ions, depending on the concentration of the reagents. Protonated species were even present in appreciable concentrations. All the found species have taurodeoxycholate aggregation numbers in multiples of three. A mechanism for the micellar aggregates containing calcium and sodium is proposed. Sodium taurodeoxycholate in the presence of calcium(II) forms larger aggregates than does taurocholate in the presence of calcium(II); the building block of the former is a trimer whereas the latter system has lower aggregation numbers and its building block is a dimer or an octamer.     

Sodium/calcium selectivity of cloned calcium T-type channels

We studied the peculiarities of permeability with respect to the main extracellular cations, Na⁺ and Ca²⁺, of cloned low-threshold calcium channels (LTCCs) of three subtypes, Ca_v3.1 (α1G), Ca_v3.2 (α 1H), and Ca_v3.3 (α1I), functionally expressed in Xenopus oocytes. In a calcium-free solution containing 100 mM Na⁺ and 5 mM calcium-chelating EGTA buffer (to eliminate residual concentrations of Ca²⁺) we observed considerable integral currents possessing the kinetics of inactivation typical of LTCCs and characterized by reversion potentials of −10 ± 1, −12 ± 1, and −18 ± 2 mV, respectively, for Ca_v3.1, Ca_v3.2, and Ca_v3.3 channels. The presence of Ca²⁺ in the extracellular solution exerted an ambiguous effect on the examined currents. On the one hand, Ca²⁺ effectively blocked the current of monovalent cations through cloned LTCCs (K _d = 2, 10, and 18 μM for currents through channels Ca_v3.1, Ca_v3.2, and Ca_v3.3, respectively). On the other hand, at the concentration of 1 to 100 mM, Ca²⁺ itself functioned as a carrier of the inward current. Despite the fact that the calcium current reached the level of saturation in the presence of 5 mM Ca²⁺ in the external solution, extracellular Na⁺ influenced the permeability of these channels even in the presence of 10 mM Ca²⁺. The Ca_v3.3 channels were more permeable with respect to Na⁺ (P _Ca/P _Na ∼ 21) than Ca_v3.1 and Ca_v3.2 (P _Ca/P _Na ∼ 66). As a whole, our data indicate that cloned LTCCs form multi-ion Ca²⁺-selective pores, as these ions possess a high affinity for certain binding sites. Monovalent cations present together with Ca²⁺ in the external solution modulate the calcium permeability of these channels. Among the above-mentioned subtypes, Ca_v3.3 channels show the minimum selectivity with respect to Ca²⁺ and are most permeable for monovalent cations. Neirofiziologiya/Neurophysiology, Vol. 38, No. 3, pp. 183–192, May–June, 2006.     

Studies on the mechanism of Salmonella typhimurium enterotoxin-induced diarrhoea

The unidirectional fluxes of Na⁺ and Cl^? were studied in Salmonella typhimurium enterotoxin-treated rats. There was net secretion of Na⁺ and Cl^? in toxin-treated animals, while in control animals there was net absorption of these ions. In the presence of the Ca²⁺-ionophore, there was net secretion of Na²⁺ and Cl^? in the control group, while the ionophore enhanced the secretion of these ions in experimental anaimals. The calcium channel blocker, verapamil, decreased the secretion induced by salmonella toxin, but could not reverse the secretion of absorption. There was no difference in the net absorption of Ca²⁺ in both the control and experimental animals. There was a significant increase in the intracellular free calcium concentrations in enterocytes isolated from toxin-treated rat intestines as compared to that in enterocytes isolated from control animals. In the presence of PMA (phorobol-12-myristated-13-acetate) there was net secretion of Na⁺ and Cl^? in the control group, while in the experimental group there was no change in the fluxes of these ions. The selective, potent inhibitor of protein kinase C, H-7 (1-(5-isoquinolinylsulphonyl)-2-methylpiperazine)_reversed the secretion of Na⁺ and Cl^? in the toxin-treated group to absorption. The addition of indomethacin also inhibited the secretion induced by salmonella toxin, but failed to reverse it to absorption. However, the addition both H-7 and indomethacin to the experimental group had a partial additive effect. These studies demonstrate that the Salmonella enterotoxin-mediated fluid secretion involved protein kinase C and the arachidonic acid metabolites and perhaps does not involve the extracellular calcium pools.     

Active transport of Calcium ions in the skin of the salamander Ambystoma tigrinum

We measured Ca²⁺ exchanges across the skin of larval and adult Ambystoma tigrinum using the radio-isotope influx method. We found that the skin of both morphs takes up Ca²⁺ in a manner that is proportional to external [Ca²⁺], saturable and oriented against the electrochemical gradient for Ca²⁺. We conclude that this uptake occurs by active transport. Kinetic analysis yields affinities for calcium ions that are similar to the affinities for both Ca²⁺ and Na⁺ in the skin of other amphibians. The capacity for calcium is similar to Ca²⁺ capacity in other amphibians. The capacity for Ca²⁺ is lower than the capacity for Na⁺. Cutaneous Ca²⁺ deposits are lower in this urodele than found in anurans. Adults tend to have higher levels of Ca in their skin than do larvae. Accepted: 25 June 1999     

The ionic hemolymph composition of the terrestrial isopod Porcellio scaber Latr. during molt

We analyzed the ionic composition of the hemolymph of Porcellio scaber in four different stages of the molt cycle using capillary electrophoresis and calcium selective mini- and microelectrodes. The main ions in the hemolymph were K⁺, Ca²⁺, Na⁺, Mg⁺, and Cl⁻. The values for total calcium obtained by means of capillary electrophoresis and calcium selective minielectrodes did not differ significantly from each other. In situ measurements of the free Ca²⁺ concentration ([Ca²⁺]) by means of calcium-selective microelectrodes indicated that Ca²⁺ is not bound in the hemolymph. During molt the [Ca²⁺] is significantly larger than during intermolt. The [Ca²⁺] increased by 13%, 19% and 18% during premolt, intramolt, and postmolt, respectively. The concentration of the other cations and of Cl⁻ decreased significantly between premolt and intramolt. Thus, the rise of the [Ca²⁺] in the hemolymph is not due to a general increase in all ions, but rather to the resorption of cuticular calcium. Furthermore, the results suggest that K⁺, Na⁺, Mg⁺, and Cl⁻are extruded from the hemolymph during and/or after posterior ecdysis. Accepted: 5 August 1997     

On the special role of NCX in astrocytes: Translating Na+-transients into intracellular Ca2+ signals

As a solute carrier electrogenic transporter, the sodium/calcium exchanger (NCX1-3/SLC8A1-A3) links the trans-plasmalemmal gradients of sodium and calcium ions (Na⁺, Ca²⁺) to the membrane potential of astrocytes. Classically, NCX is considered to serve the export of Ca²⁺ at the expense of the Na⁺ gradient, defined as a “forward mode” operation. Forward mode NCX activity contributes to Ca²⁺ extrusion and thus to the recovery from intracellular Ca²⁺ signals in astrocytes. The reversal potential of the NCX, owing to its transport stoichiometry of 3 Na⁺ to 1 Ca²⁺, is, however, close to the astrocytes’ membrane potential and hence even small elevations in the astrocytic Na⁺ concentration or minor depolarisations switch it into the “reverse mode” (Ca²⁺ import/Na⁺ export). Notably, transient Na⁺ elevations in the millimolar range are induced by uptake of glutamate or GABA into astrocytes and/or by the opening of Na⁺-permeable ion channels in response to neuronal activity. Activity-related Na⁺ transients result in NCX reversal, which mediates Ca²⁺ influx from the extracellular space, thereby generating astrocyte Ca²⁺ signalling independent from InsP₃-mediated release from intracellular stores. Under pathological conditions, reverse NCX promotes cytosolic Ca²⁺ overload, while dampening Na⁺ elevations of astrocytes. This review provides an overview on our current knowledge about this fascinating transporter and its special functional role in astrocytes. We shall delineate that Na⁺-driven, reverse NCX-mediated astrocyte Ca²⁺ signals are involved neurone-glia interaction. Na⁺ transients, translated by the NCX into Ca²⁺ elevations, thereby emerge as a new signalling pathway in astrocytes.     

Ionic selectivity in L-type calcium channels by electrostatics and hard-core repulsion

A physical model of selective “ion binding” in the L-type calcium channel is constructed, and consequences of the model are compared with experimental data. This reduced model treats only ions and the carboxylate oxygens of the EEEE locus explicitly and restricts interactions to hard-core repulsion and ion–ion and ion–dielectric electrostatic forces. The structural atoms provide a flexible environment for passing cations, thus resulting in a self-organized induced-fit model of the selectivity filter. Experimental conditions involving binary mixtures of alkali and/or alkaline earth metal ions are computed using equilibrium Monte Carlo simulations in the grand canonical ensemble. The model pore rejects alkali metal ions in the presence of biological concentrations of Ca²⁺ and predicts the blockade of alkali metal ion currents by micromolar Ca²⁺. Conductance patterns observed in varied mixtures containing Na⁺ and Li⁺, or Ba²⁺ and Ca²⁺, are predicted. Ca²⁺ is substantially more potent in blocking Na⁺ current than Ba²⁺. In apparent contrast to experiments using buffered Ca²⁺ solutions, the predicted potency of Ca²⁺ in blocking alkali metal ion currents depends on the species and concentration of the alkali metal ion, as is expected if these ions compete with Ca²⁺ for the pore. These experiments depend on the problematic estimation of Ca²⁺ activity in solutions buffered for Ca²⁺ and pH in a varying background of bulk salt. Simulations of Ca²⁺ distribution with the model pore bathed in solutions containing a varied amount of Li⁺ reveal a “barrier and well” pattern. The entry/exit barrier for Ca²⁺ is strongly modulated by the Li⁺ concentration of the bath, suggesting a physical explanation for observed kinetic phenomena. Our simulations show that the selectivity of L-type calcium channels can arise from an interplay of electrostatic and hard-core repulsion forces among ions and a few crucial channel atoms. The reduced system selects for the cation that delivers the largest charge in the smallest ion volume.     

Requirement of monovalent cations in the acrosome reaction of guinea pig spermatozoa

The majority of the spermatozoa precapacitated in Ca²⁺-free medium underwent the acrosome raction rapidly when they were transferred to Ca²⁺-containing medium. The presence of Na⁺ and Ca²⁺ in the medium was essential for the acrosome reaction. The vast majority of spermatozoa failed to undergo the reaction in Ca²⁺ medium lacking monovalent ions, although they remained motile. At the concentration of 140 mM, Na⁺, K⁺, Rb⁺, and Cs⁺ all supported the reaction at the maximum level, but at 50 mM the latter three ions were not as effective as Na⁺. Li⁺ was least effective in supporting the reaction. Virtually no acrosome reactions took place when precapacitated spermatozoa were first exposed to Na⁺ medium (no Ca²⁺) and then to Ca²⁺ medium (no Na⁺). On the other hand, a considerably higher proportion of spermatozoa acrosome reacted when they were exposed to these media in the reverse order. The most efficient acrosome reactions took place when the medium contained both a monovalent ion (Na⁺) and Ca²⁺ simultaneously. Possible mechanisms by which monovalent and divalent cations participate in the acrosome reaction are discussed.     

Response of red-osier dogwood (Cornus stolonifera) seedlings to sodium sulphate salinity: effects of supplemental calcium

A greenhouse study was designed to test the effects of sodium sulphate (Na₂SO₄) on red-osier dogwood (Cornus stolonifera Michx) seedlings in the presence and absence of additional calcium (Ca²⁺). Changes in growth parameters, ion and carbohydrate accumulation and cell wall composition were examined. Calcium alleviated the effect of Na₂SO₄ on shoot height; however, it did not affect the reduction in shoot and root dry weights. An increased level of sodium (Na⁺) in roots of plant exposed to Na₂SO₄ was recorded in the presence of supplemental Ca²⁺ whereas there was no change in potassium (K⁺) and Ca²⁺ levels. In shoots of seedlings treated with Na₂SO₄, the addition of Ca²⁺ did not affect Na⁺, K⁺ and Ca²⁺ levels. The amount of soluble carbohydrates was increased in leaves of seedlings treated with Na₂SO₄ both in the absence and presence of supplemental Ca²⁺. The decrease in cell wall material in response to salt stress was alleviated by Ca²⁺ in stem tissues although Ca²⁺ did not alter the changes in hemicellulose and cellulose. Sugar composition of pectins and hemicellulose were modified in stems and leaves by Na₂SO₄ and/or Ca²⁺. The results of this study showed that calcium was able to alleviate the effects of salt stress on shoot height and cell wall content of red-osier dogwood stems. Furthermore, changes occurred in cell wall composition of red osier seedlings treated with Na₂SO₄.     

Sodium/calcium exchanger in rat olfactory neurons

The chemo-electrical transduction process in olfactory neurons is accompanied by a rapid and transient increase in intracellular calcium concentrations. The notion that Na⁺/Ca²⁺ exchanger activities may play a major role in extruding calcium ions out of the cell and maintaining Ca²⁺ homeostasis in olfactory receptor cells was assessed by means of laser scanning confocal microscopy in combination with the fluorescent indicators Fluo-3 and Fura-Red. The data indicate that high exchanger acitivity, which was inhibited by amiloride derivatives, is located in the dendritic knob and probably in the olfactory cilia. This result was supported by experiments using specific antiserum raised against retinal Na⁺/Ca²⁺ exchanger protein which labelled an immunoreactive protein of 230 kDa in Western blots from olfactory tissue and strongly stained the ciliary layer of the olfactory epithelium.     

Reduced external calcium or sodium stimulates calcium influx in Pelvetia eggs

The effect of external calcium and sodium ion concentrations on the calcium fluxes on the Pelvetia fastigiata De Toni egg was measured. Decreasing external [Ca²⁺] greatly increased the permeability of the eggs to Ca²⁺; at 1 mM external Ca²⁺ this permeability was 60 times as great as it was at the normal [Ca²⁺] of 10 mM. Lowering the external [Na⁺] also increased Ca²⁺ influx; at 2 mM Na⁺, the Ca²⁺ influx was 2–3 times as great as it was at the normal [Na⁺] if choline was used as a Na⁺ substitute. Lithium was less effective as a Na⁺ substitute in increasing Ca²⁺ influx. The extra Ca²⁺ influx in low [Na⁺] seemed to be dependent on internal [Na⁺]. The Ca²⁺ efflux increased transiently and then declined in low Na⁺ media.     

Nuclear localization of NCX: Role in Ca2+ handling and pathophysiological implications

Numerous lines of evidence indicate that nuclear calcium concentration ([Ca²⁺]_n) may be controlled independently from cytosolic events by a local machinery. In particular, the perinuclear space between the inner nuclear membrane (INM) and the outer nuclear membrane (ONM) of the nuclear envelope (NE) likely serves as an intracellular store for Ca²⁺ ions. Since ONM is contiguous with the endoplasmic reticulum (ER), the perinuclear space is adjacent to the lumen of ER thus allowing a direct exchange of ions and factors between the two organelles. Moreover, INM and ONM are fused at the nuclear pore complex (NPC), which provides the only direct passageway between the nucleoplasm and cytoplasm. However, due to the presence of ion channels, exchangers and transporters, it has been generally accepted that nuclear ion fluxes may occur across ONM and INM. Within the INM, the Na⁺/Ca²⁺ exchanger (NCX) isoform 1 seems to play an important role in handling Ca²⁺ through the different nuclear compartments. Particularly, nuclear NCX preferentially allows local Ca²⁺ flowing from nucleoplasm into NE lumen thanks to the Na⁺ gradient created by the juxtaposed Na⁺/K⁺-ATPase. Such transfer reduces abnormal elevation of [Ca²⁺]_n within the nucleoplasm thus modulating specific transductional pathways and providing a protective mechanism against cell death. Despite very few studies on this issue, here we discuss those making major contribution to the field, also addressing the pathophysiological implication of nuclear NCX malfunction.     

A comparative analysis of the contracture responses induced by acetylcholine and choline in the twitch and tonic fibers of frog skeletal muscles

A comparative analysis of the contractile responses induced by acetylcholine and replacement of the external Na⁺ ions with choline ions in the isolated twitch and tonic fibers of frog skeletal muscles was performed. The effects of extracellular Ca²⁺ concentration and several pharmacological agents modulating the activity of various systems maintaining Ca²⁺ level in the myoplasm (dantrolene, cresol, d-tubocurarine, and tetrodotoxin) were studied. It has been found that a voltage-dependent Ca²⁺ release from the sarcoplasmic reticulum depot is the main mechanism inducing the acetylcholine contracture in the fibers of both types. However, the twitch and tonic fibers differ in the properties of the α-isoform and(or) the ratio of α- to β-isoforms of ryanodine-sensitive channels. In the fibers of both types, the replacement of over 25% of Na⁺ ions with choline induces long-term contracture responses, which are also mediated by activation of acetylcholine receptors. It is assumed that an additional mechanism—accumulation of choline ions in the myoplasm and their direct action on the ryanodine-sensitive channels—is involved in the development of such contractile responses.