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.     

Na+−Ca2+ exchanger: From basics to molecular biology

The plasmalemmal Na⁺−Ca²⁺ exchanger is a coupled Na⁺ and Ca²⁺ transport mechanism that plays an important role in regulation of Ca²⁺ homeoslasis in many cell types. A robust Na⁺−Ca²⁺ exchange system is present in the heart where it comprises essential Ca²⁺ extrusion, as well as Ca²⁺ entry pathways, that significantly contribute to the maintenance of cardiac contractility. The review examines the basic properties of Na⁺−Ca²⁺ exchange, the patterns of its regulation, as well as the latest achievements in the cloning and structure-function studies of a Na⁺−Ca²⁺ exchanger molecule.     

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.     

Function and Regulation of the Na+-Ca2+ Exchanger NCX3 Splice Variants in Brain and Skeletal Muscle

Isoform 3 of the Na⁺-Ca²⁺ exchanger (NCX3) is crucial for maintaining intracellular calcium ([Ca²⁺]_i) homeostasis in excitable tissues. In this sense NCX3 plays a key role in neuronal excitotoxicity and Ca²⁺ extrusion during skeletal muscle relaxation. Alternative splicing generates two variants (NCX3-AC and NCX3-B). Here, we demonstrated that NCX3 variants display a tissue-specific distribution in mice, with NCX3-B as mostly expressed in brain and NCX-AC as predominant in skeletal muscle. Using Fura-2-based Ca²⁺ imaging, we measured the capacity and regulation of the two variants during Ca²⁺ extrusion and uptake in different conditions. Functional studies revealed that, although both variants are activated by intracellular sodium ([Na⁺]_i), NCX3-AC has a higher [Na⁺]_i sensitivity, as Ca²⁺ influx is observed in the presence of extracellular Na⁺. This effect could be partially mimicked for NCX3-B by mutating several glutamate residues in its cytoplasmic loop. In addition, NCX3-AC displayed a higher capacity of both Ca²⁺ extrusion and uptake compared with NCX3-B, together with an increased sensitivity to intracellular Ca²⁺. Strikingly, substitution of Glu⁵⁸⁰ in NCX3-B with its NCX3-AC equivalent Lys⁵⁸⁰ recapitulated the functional properties of NCX3-AC regarding Ca²⁺ sensitivity, Lys⁵⁸⁰ presumably acting through a structure stabilization of the Ca²⁺ binding site. The higher Ca²⁺ uptake capacity of NCX3-AC compared with NCX3-B is in line with the necessity to restore Ca²⁺ levels in the sarcoplasmic reticulum during prolonged exercise. The latter result, consistent with the high expression in the slow-twitch muscle, suggests that this variant may contribute to the Ca²⁺ handling beyond that of extruding Ca²⁺.     

Modulation of calcium signalling by mitochondria

In this review we will attempt to summarise the complex and sometimes contradictory effects that mitochondria have on different forms of calcium signalling. Mitochondria can influence Ca²⁺ signalling indirectly by changing the concentration of ATP, NAD(P)H, pyruvate and reactive oxygen species — which in turn modulate components of the Ca²⁺ signalling machinery i.e. buffering, release from internal stores, influx from the extracellular solution, uptake into cellular organelles and extrusion by plasma membrane Ca²⁺ pumps. Mitochondria can directly influence the calcium concentration in the cytosol of the cell by importing Ca²⁺ via the mitochondrial Ca²⁺ uniporter or transporting Ca²⁺ from the interior of the organelle into the cytosol by means of Na⁺/Ca²⁺ or H⁺/Ca²⁺ exchangers. Considerable progress in understanding the relationship between Ca²⁺ signalling cascades and mitochondrial physiology has been accumulated over the last few years due to the development of more advanced optical techniques and electrophysiological approaches.     

Purinergic stimulation of K+-dependent Na+/Ca2+ exchanger isoform 4 requires dual activation by PKC and CaMKII

K⁺-dependent Na⁺/Ca²⁺-exchanger isoform 4 (NCXK4) is one of the most broadly expressed members of the NCKX (K⁺-dependent Na⁺/Ca²⁺-exchanger) family. Recent data indicate that NCKX4 plays a critical role in controlling normal Ca²⁺ signal dynamics in olfactory and other neurons. Synaptic Ca²⁺ dynamics are modulated by purinergic regulation, mediated by ATP released from synaptic vesicles or from neighbouring glial cells. Previous studies have focused on modulation of Ca²⁺ entry pathways that initiate signalling. Here we have investigated purinergic regulation of NCKX4, a powerful extrusion pathway that assists in terminating Ca²⁺ signals. NCKX4 activity was stimulated by ATP through activation of the P2Y receptor signalling pathway. Stimulation required dual activation of PKC (protein kinase C) and CaMKII (Ca²⁺/calmodulin-dependent protein kinase II). Mutating T312, a putative PKC phosphorylation site on NCKX4, partially prevented purinergic stimulation. These data illustrate how purinergic regulation can shape the dynamics of Ca²⁺ signalling by activating a signal damping and termination pathway.     

Calcium extrusion mechanisms in dendrites of mouse hippocampal CA1 inhibitory interneurons

Local circuit GABAergic inhibitory interneurons control the integration and transfer of information in many brain regions. Several different forms of plasticity reported at interneuron excitatory synapses are triggered by cell- and synapse-specific postsynaptic calcium (Ca²⁺) mechanisms. To support this function, the spatiotemporal dynamics of dendritic Ca²⁺ elevations must be tightly regulated. While the dynamics of postsynaptic Ca²⁺ signaling through activation of different Ca²⁺ sources has been explored, the Ca²⁺ extrusion mechanisms that operate in interneuron dendrites during different patterns of activity remain largely unknown. Using a combination of whole-cell patch-clamp recordings and two-photon Ca²⁺ imaging in acute mouse hippocampal slices, we characterized the Ca²⁺ extrusion mechanisms activated by Ca²⁺ transients (CaTs) associated with backpropagating action potentials (bAPs) in dendrites of hippocampal CA1 stratum radiatum interneurons. Our data showed that Ca²⁺ clearance increased as a function of activity, pointing to an activity-dependent recruitment of specific Ca²⁺ extrusion mechanisms. bAP-CaTs were significantly prolonged in the presence of the plasma membrane Ca²⁺ ATPase (PMCA) and Na⁺/Ca²⁺ exchanger (NCX) inhibitors as well as the sarco/endoplasmic reticulum Ca²⁺ ATPase (SERCA) and the mitochondria Ca²⁺ uniporter (MCU) blockers. While PMCA, NCX and SERCA pumps cooperated in the cytosolic Ca²⁺ removal at a wide range of concentrations, the MCU was only activated at higher Ca²⁺ loads produced by repetitive interneuron firing. These results identify a division of labor between distinct Ca²⁺ extrusion mechanisms shaping dendritic Ca²⁺ dynamics and possibly contributing to activity-dependent regulation of synaptic inputs in interneurons. In addition, the MCU activated by larger Ca²⁺ levels may be involved in the activity-dependent ATP production or interneuron-selective vulnerability associated with cytosolic Ca²⁺ overloads under pathological conditions.     

Mechanisms underlying odorant-induced and spontaneous calcium signals in olfactory receptor neurons of spiny lobsters, Panulirus argus

We determined if a newly developed antennule slice preparation allows studying chemosensory properties of spiny lobster olfactory receptor neurons under in situ conditions with Ca²⁺ imaging. We show that chemical stimuli reach the dendrites of olfactory receptor neurons but not their somata, and that odorant-induced Ca²⁺ signals in the somata are sufficiently stable over time to allow stimulation with a substantial number of odorants. Pharmacological manipulations served to elucidate the source of odorant-induced Ca²⁺ transients and spontaneous Ca²⁺ oscillations in the somata of olfactory receptor neurons. Both Ca²⁺ signals are primarily mediated by an influx of extracellular Ca²⁺ through voltage-activated Ca²⁺ channels that can be blocked by CoCl₂ and the L-type Ca²⁺ channel blocker verapamil. Intracellular Ca²⁺ stores contribute little to odorant-induced Ca²⁺ transients and spontaneous Ca²⁺ oscillations. The odorant-induced Ca²⁺ transients as well as the spontaneous Ca²⁺ oscillations depend on action potentials mediated by Na⁺ channels that are largely TTX-insensitive but blocked by the local anesthetics tetracaine and lidocaine. Collectively, these results corroborate the conclusion that odorant-induced Ca²⁺ transients and spontaneous Ca²⁺ oscillations in the somata of olfactory receptor neurons closely reflect action potential activity associated with odorant-induced phasic-tonic responses and spontaneous bursting, respectively. Therefore, both types of Ca²⁺ signals represent experimentally accessible proxies of spiking.     

Na+-dependent Ca2+ Extrusion Governs Response Recovery in Frog Olfactory Receptor Cells

To study the mechanism by which Ca²⁺, which enters during the odor response, is extruded during response recovery, recordings were made from isolated frog olfactory receptor cells using the suction pipette technique, while superfusing the olfactory cilia with solutions of modified ionic composition. When external Na⁺ was substituted with another cation, the response to odor was greatly prolonged. This prolongation of the response was similar irrespective of whether Na⁺ was replaced with Li⁺, which permeates the cyclic nucleotide-gated conductance, or choline, which does not. The prolonged current was greatly reduced by exposure to 300 μM niflumic acid, a blocker of the calcium-activated chloride channel, indicating that it is carried by this conductance, and abolished if Ca²⁺ was omitted from the external solution, demonstrating that Ca²⁺ influx is required for its generation. When the cilia were exposed to Na⁺-free solution after odor stimulation, the recovery of the response to a second stimulus from the adaptation induced by the first was greatly reduced. We conclude that a Na⁺-dependent Ca²⁺ extrusion mechanism is present in frog olfactory cilia and that it serves as the main mechanism that returns cytoplasmic Ca²⁺ concentration to basal levels after stimulation and mediates the normally rapid recovery of the odor response and the restoration of sensitivity after adaptation.     

Oxygen free radicals and calcium homeostasis in the heart

Many experiments have been done to clarify the effects of oxygen free radicals on Ca²⁺ homeostasis in the hearts. A burst of oxygen free radicals occurs immediately after reperfusion, but we have to be reminded that the exact levels of oxygen free radicals in the hearts are yet unknown in both physiological and pathophysiological conditions. Therefore, we should give careful consideration to this point when we perform the experiments and analyze the results. It is, however, evident that Ca²⁺ overload occurs when the hearts are exposed to an excess amount of oxygen free radicals. Though ATP-independent Ca²⁺ binding is increased, Ca²⁺ influx through Ca²⁺ channel does not increase in the presence of oxygen free radicals. Another possible pathway through which Ca²⁺ can enter the myocytes is Na⁺?Ca²⁺ exchanger. Although, the activities of Na⁺?K⁺ ATPase and Na⁺?H⁺ exchange are inhibited by oxygen free radicals, it is not known whether intracellular Na⁺ level increases under oxidative stress or not. The question has to be solved for the understanding of the importance of Na⁺?Ca²⁺ exchange in Ca²⁺ influx process from extracellular space. Another question is ‘which way does Na⁺?Ca²⁺ exchange work under oxidative stress? Net influx or efflux of Ca^2+?’ Membrane permeability for Ca²⁺ may be maintained in a relatively early phase of free radical injury. Since sarcolemmal Ca²⁺-pump ATPase activity is depressed by oxygen free radicals, Ca²⁺ extrusion from cytosol to extracellular space is considered to be reduced. It has also been shown that oxygen free radicals promote Ca²⁺ release from sarcoplasmic reticulum and inhibit Ca²⁺ sequestration to sarcoplasmic reticulum. Thus, these changes in Ca²⁺ handling systems could cause the Ca²⁺ overload due to oxygen free radicals.     

Olfactory response termination involves Ca2+-ATPase in vertebrate olfactory receptor neuron cilia

In vertebrate olfactory receptor neurons (ORNs), odorant-induced activation of the transduction cascade culminates in production of cyclic AMP, which opens cyclic nucleotide–gated channels in the ciliary membrane enabling Ca²⁺ influx. The ensuing elevation of the intraciliary Ca²⁺ concentration opens Ca²⁺-activated Cl⁻ channels, which mediate an excitatory Cl⁻ efflux from the cilia. In order for the response to terminate, the Cl⁻ channel must close, which requires that the intraciliary Ca²⁺ concentration return to basal levels. Hitherto, the extrusion of Ca²⁺ from the cilia has been thought to depend principally on a Na⁺–Ca²⁺ exchanger.In this study, we show using simultaneous suction pipette recording and Ca²⁺-sensitive dye fluorescence measurements that in fire salamander ORNs, withdrawal of external Na⁺ from the solution bathing the cilia, which incapacitates Na⁺–Ca²⁺exchange, has only a modest effect on the recovery of the electrical response and the accompanying decay of intraciliary Ca²⁺ concentration. In contrast, exposure of the cilia to vanadate or carboxyeosin, a manipulation designed to block Ca²⁺-ATPase, has a substantial effect on response recovery kinetics. Therefore, we conclude that Ca²⁺-ATPase contributes to Ca²⁺ extrusion in ORNs, and that Na⁺–Ca²⁺exchange makes only a modest contribution to Ca²⁺ homeostasis in this species.     

An inhibitor of Na/Ca exchange blocks activation of insect olfactory receptors

Earlier we showed that the Na⁺/Ca²⁺ exchanger inhibitor, KB-R7943, potently blocks the odor-evoked activity of lobster olfactory receptor neurons. Here we extend that finding to recombinant mosquito olfactory receptors stably expressed in HEK cells. Using whole-cell and outside-out patch clamping and calcium imaging, we demonstrate that KB-R7943 blocks both the odorant-gated current and the odorant-evoked calcium signal from two different OR complexes from the malaria vector mosquito, Anopheles gambiae, AgOr48 + AgOrco and AgOr65 + AgOrco. Both heteromeric and homomeric (Orco alone) OR complexes were susceptible to KB-R7943 blockade when activated by VUAA1, an agonist that targets the Orco channel subunit, suggesting the Orco subunit may be the target of the drug’s action. KB-R7943 represents a valuable tool to further investigate the functional properties of arthropod olfactory receptors and raises the interesting specter that activation of these ionotropic receptors is directly or indirectly linked to a Na⁺/Ca²⁺ exchanger, thereby providing a template for drug design potentially allowing improved control of insect pests and disease vectors.     

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.     

Ouabain-insensitive Na+-stimulated ATPase activity of basolateral plasma membranes from guinea-pig kidney cortex cells: II. Effect of Ca2+

The ouabain-insensitive, Mg²⁺-dependent, Na⁺-stimulated ATPase activity present in fresh basolateral plasma membranes from guinea-pig kidney cortex cells (prepared at pH 7.2) can be increased by the addition of micromolar concentrations of Ca²⁺ to the assay medium. The Ca²⁺ involved in this effect seems to be associated with the membranes in two different ways: as a labile component, which can be quickly and easily ‘deactivated’ by reducing the free Ca²⁺ concentration of the assay medium to values lower than 1 μM; and as a stable component, which can be ‘deactivated’ by preincubating the membranes for periods of 3–4 h with 2 mM EDTA or EGTA. Both components are easily activated by micromolar concentrations of Ca²⁺. The $\text{K}{}_{\mathrm{<mtext>a</mtext>}}$ of the system for Na⁺ is the same, 8 mM, whether only the stable component or both components, stable and labile, are working. In other words, the activating effect of Ca²⁺ on the Na⁺-stimulated ATPase is on the $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$, and not on the $\text{K}{}_{\mathrm{<mtext>a</mtext>}}$ of the system for Na⁺. The activating effect of Ca²⁺ may be related to some conformational change produced by the interaction of this ion with the membranes, since it can also be obtained by resuspending the membranes at pH 7.8 or by ageing the preparations. Changes in the Ca²⁺ concentration may modulate the ouabain-insensitive, Na⁺-stimulated ATPase activity. This modulation could regulate the magnitude of the extrusion of Na⁺ accompanied by Cl^? and water that these cells show, and to which the Na⁺-ATPase has been associated as being responsible for the energy supply of this mode of Na⁺ extrusion.     

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.     

NCLX Protein,but Not LETM1, Mediates Mitochondrial Ca2+ Extrusion,Thereby Limiting Ca2+-induced NAD(P)H Production and Modulating Matrix Redox State

Mitochondria capture and subsequently release Ca²⁺ ions, thereby sensing and shaping cellular Ca²⁺ signals. The Ca²⁺ uniporter MCU mediates Ca²⁺ uptake, whereas NCLX (mitochondrial Na/Ca exchanger) and LETM1 (leucine zipper-EF-hand-containing transmembrane protein 1) were proposed to exchange Ca²⁺ against Na⁺ or H⁺, respectively. Here we study the role of these ion exchangers in mitochondrial Ca²⁺ extrusion and in Ca²⁺-metabolic coupling. Both NCLX and LETM1 proteins were expressed in HeLa cells mitochondria. The rate of mitochondrial Ca²⁺ efflux, measured with a genetically encoded indicator during agonist stimulations, increased with the amplitude of mitochondrial Ca²⁺ ([Ca²⁺]_mt) elevations. NCLX overexpression enhanced the rates of Ca²⁺ efflux, whereas increasing LETM1 levels had no impact on Ca²⁺ extrusion. The fluorescence of the redox-sensitive probe roGFP increased during [Ca²⁺]_mt elevations, indicating a net reduction of the matrix. This redox response was abolished by NCLX overexpression and restored by the Na⁺/Ca²⁺ exchanger inhibitor {"type":"entrez-protein","attrs":{"text":"CGP37157","term_id":"875406365","term_text":"CGP37157"}}CGP37157. The [Ca²⁺]_mt elevations were associated with increases in the autofluorescence of NAD(P)H, whose amplitude was strongly reduced by NCLX overexpression, an effect reverted by Na⁺/Ca²⁺ exchange inhibition. We conclude that NCLX, but not LETM1, mediates Ca²⁺ extrusion from mitochondria. By controlling the duration of matrix Ca²⁺ elevations, NCLX contributes to the regulation of NAD(P)H production and to the conversion of Ca²⁺ signals into redox changes.     

Functional characterization of two distinct Mg2+ extrusion mechanisms in cardiac sarcolemmal vesicles

Cardiac ventricular myocytes extrude a sizeable amount of their total Mg²⁺ content upon stimulation by β-adrenergic agonists. This extrusion occurs within a few minutes from the application of the agonist, suggesting the operation of rapid and abundantly represented Mg²⁺ transport mechanisms in the cardiac sarcolemma. The present study was aimed at characterizing the operation of these transport mechanisms under well defined conditions. Male Sprague-Dawley rats were used to purify a biochemical standardized preparation of sealed rat cardiac sarcolemmal vesicles. This experimental model has the advantage that trans-sarcolemmal cation transport can be studied under specific extra- and intra-vesicular ionic conditions, in the absence of intracellular organelles, and buffering or signaling components. Magnesium ion (Mg²⁺) transport was assessed by atomic absorbance spectrophotometry. The results reported here indicate that: (1) sarcolemma vesicles retained trapped intravesicular Mg²⁺ in the absence of extravesicular counter-ions; (2) the addition of Na⁺ or Ca²⁺ induced a rapid and concentration-dependent Mg²⁺ extrusion from the vesicles; (3) co-addition of maximal concentrations of Na⁺ and Ca²⁺ resulted in an additive Mg²⁺ extrusion; (4) Mg²⁺ extrusion was blocked by addition of amiloride or imipramine; (5) pre-treatment of sarcolemma vesicles with alkaline phosphatase at the time of preparation completely abolished Na⁺- but not Ca²⁺-induced Mg²⁺ extrusion; (6) Na⁺-dependent Mg²⁺ transport could be restored by stimulating vesicles loaded with protein kinase A catalytic subunit and ATP with membrane-permeant cyclic-AMP analog; (7) extra-vesicular Mg²⁺ could be accumulated in exchange for intravesicular Na⁺ via a mechanism inhibited by amiloride or alkaline phosphatase treatment; (8) Mg²⁺ accumulation could be restored via cAMP/protein kinase A protocol. Overall, these data provide compelling evidence for the operation of distinct Na⁺- and Ca²⁺-dependent Mg²⁺ extrusion mechanisms in sarcolemma vesicles. The Na⁺-dependent mechanism appears to be specifically activated via protein kinase A/cAMP-dependent phosphorylation process, and can operate in either direction based upon the cation concentration gradient across the sarcolemma. The Ca²⁺-dependent mechanism, instead, only mediates Mg²⁺ extrusion in a cAMP-independent manner.     

Analysis of the Na+/Ca2+ Exchanger Gene Family within the Phylum Nematoda

Na⁺/Ca²⁺ exchangers are low affinity, high capacity transporters that rapidly transport calcium at the plasma membrane, mitochondrion, endoplasmic (and sarcoplasmic) reticulum, and the nucleus. Na⁺/Ca²⁺ exchangers are widely expressed in diverse cell types where they contribute homeostatic balance to calcium levels. In animals, Na⁺/Ca²⁺ exchangers are divided into three groups based upon stoichiometry: Na⁺/Ca²⁺ exchangers (NCX), Na⁺/Ca²⁺/K⁺ exchangers (NCKX), and Ca²⁺/Cation exchangers (CCX). In mammals there are three NCX genes, five NCKX genes and one CCX (NCLX) gene. The genome of the nematode Caenorhabditis elegans contains ten Na⁺/Ca²⁺ exchanger genes: three NCX; five CCX; and two NCKX genes. Here we set out to characterize structural and taxonomic specializations within the family of Na⁺/Ca²⁺ exchangers across the phylum Nematoda. In this analysis we identify Na⁺/Ca²⁺ exchanger genes from twelve species of nematodes and reconstruct their phylogenetic and evolutionary relationships. The most notable feature of the resulting phylogenies was the heterogeneous evolution observed within exchanger subtypes. Specifically, in the case of the CCX exchangers we did not detect members of this class in three Clade III nematodes. Within the Caenorhabditis and Pristionchus lineages we identify between three and five CCX representatives, whereas in other Clade V and also Clade IV nematode taxa we only observed a single CCX gene in each species, and in the Clade III nematode taxa that we sampled we identify NCX and NCKX encoding genes but no evidence of CCX representatives using our mining approach. We also provided re-annotation for predicted CCX gene structures from Heterorhabditis bacteriophora and Caenorhabditis japonica by RT-PCR and sequencing. Together, these findings reveal a complex picture of Na⁺/Ca²⁺ transporters in nematodes that suggest an incongruent evolutionary history of proteins that provide central control of calcium dynamics.     

Na+/Ca2+-K+ Exchangers (NCKX):Functional Properties and Physiological Roles

The most numerous Ca²⁺ extrusion protein family, in terms of distinct genes, is the SLC24 gene family of Na⁺/Ca²⁺-K⁺ exchangers (NCKX). Five distinct gene products have been identified, mostly from specific animal excitable tissues such as neurons and smooth muscle, but also in places like skin pigment epithelium, signifying that NCKX proteins may play very specific roles, related to Ca²⁺ homeostasis, in these tissues. However, progress in elucidating the specific physiological roles of NCKX proteins has been slow in coming, largely because of challenges relating to isolating the activity of these proteins in their native tissues. Herein, we provide an overview of NCKX protein functional characteristics, highlighting properties that are unique and useful as distinguishing features over other Ca²⁺ handling mechanisms. We also present the first comprehensive review of the literature concerning physiological roles of NCKX proteins.     

Ionophore-mediated Ca2+ countertransport: role of Na+, Li+ or H+ gradient

Summary In an artificial system, the ionophore A23187, which transports Ca²⁺ but not Na⁺, is able to mediate the uphill translocation of Ca²⁺ from one aqueous medium to another across an organic immiscible phase, provided that a Na⁺, Li⁺ or H⁺ gradient is imposed on the system. Therefore, in the process known as Na-Ca countertransport, the downhill influx of Na⁺ may not be necessary for causing Ca²⁺ extrusion against its electrochemical gradient.