Activation of the BK (SLO1) potassium channel by mallotoxin

Pharmacologic approaches to activate K+ channels represent an emerging strategy to regulate membrane excitability. Here we report the identification and characterization of a lipid soluble toxin, mallotoxin (rottlerin), which potently activates the large conductance voltage and Ca2+-activated K+ channel (BK) expressed in a heterologous expression system and human vascular smooth muscle cells, shifting the conductance/voltage relationship by >100 mV. Probing the mechanism of action, we discover that the BK channel can be activated in the absence of divalent cations (Ca2+, Mg2+), suggesting that the mallotoxin mechanism of action involves the voltage-dependent gating of the channel. Mallotoxin-activated channels remain incrementally sensitive to Ca2+ and beta subunits. In comparison to other small hydrophobic poisons, anesthetic agents, and protein toxins that inhibit ion channel activity, mallotoxin potently activates channel activity. In certain respects, mallotoxin acts as a BK channel beta1 subunit mimetic, preserving BK channel Ca2+ sensitivity yet adjusting the set-point for BK channel activation to a more hyperpolarized membrane potential.     

Characterisation of a high affinity Ca2+-stimulated, Mg2+-dependent ATPase in the rat parotid plasma membrane

Two Ca2+-stimulated ATPase activities have been identified in the plasma membrane of rat parotid: (a) a (Ca2+ + Mg2+)-ATPase with high affinity for free Ca2+ (apparent Km = 208 nM, Vmax = 188 nmol/min per mg) and requiring micromolar concentration of Mg2+ and (b) a (Ca2+ or Mg2+)-ATPase with relatively low affinity for free Ca2+ (K0.5 = 23 microM) or free Mg2+ (K0.5 = 26 microM). The low-affinity (Ca2+ or Mg2+)-ATPase can be maximally stimulated by Ca2+ alone or Mg2+ alone. The high-affinity (Ca2+ + Mg2+)-ATPase exhibits sigmoidal kinetics with respect to ATP concentration with K0.5 = 0.4 mM and a Hill coefficient of 1.91. It displays low substrate specificity with respect to nucleotide triphosphates. Although trifluoperazine inhibits the activity of the high affinity (Ca2+ + Mg2+)-ATPase only slightly, it inhibits the activity of the low-affinity (Ca2+ or Mg2+)-ATPase quite potently with 22 microM trifluoperazine inhibiting the enzymic activity by 50%. Vanadate, inositol 1,4,5-trisphosphate, phosphatidylinositol 4,5-bisphosphate, Na+,K+ and ouabain had no effect on the activities of both ATPases. Calmodulin added to the plasma membranes does not stimulate the activities of both ATPases. The properties of the high-affinity (Ca2+ + Mg2+)-ATPase are distinctly different from those of the previously reported Ca2+-pump activity of the rat parotid plasma membrane.     

Identification of a spontaneously active, Na+-permeable channel in guinea pig gallbladder smooth muscle

The action potential in gallbladder smooth muscle (GBSM) is caused by Ca2+ entry through voltage-dependent Ca2+ channels (VDCC), which contributes to the GBSM contractions. Action potential generation in GBSM is critically dependent on the resting membrane potential (about -50 mV), which is approximately 35 mV more positive of the K+ equilibrium potential. We hypothesized that a tonic, depolarizing conductance is present in GBSM and contributes to the regulation of the resting membrane potential and action potential frequency. GBSM cells were isolated from guinea pig gallbladders, and the whole cell patch-camp technique was used to record membrane currents. After eliminating the contribution of VDCC and K+ channels, we identified a novel spontaneously active cation conductance (I(cat)) in GBSM. This I(cat) was mediated predominantly by influx of Na+. Na+ substitution with N-methyl-D-glucamine (NMDG), a large relatively impermeant cation, caused a negative shift in the reversal potential of the ramp current and reduced the amplitude of the inward current at -50 mV by 65%. Membrane potential recordings with intracellular microelectrodes or in current-clamp mode of the patch-clamp technique indicated that the inhibition of I(cat) conductance by NMDG is associated with membrane hyperpolarization and inhibition of action potentials. Extracellular Ca2+, Mg2+, and Gd3+ attenuated the I(cat) in GBSM. Muscarinic stimulation did not activate the I(cat). Our results indicate that, in GBSM, an Na+-permeable channel contributes to the maintenance of the resting membrane potential and action potential generation and therefore plays a critical role in the regulation of GBSM excitability and contractility.     

A calmodulin dependent Ca2+-activated K+ channel in the adipocyte plasma membrane

Increased membrane permeability (conductance) that is specific for K+ and directly activated by Ca2+ ions, has been identified in isolated adipocyte plasma membranes using the K+ analogue, 86Rb+. Activation of these K+ conductance pathways (channels) by free Ca2+ was concentration dependent with a half-maximal effect occurring at 32 +/- 4 nM free Ca2+ (n = 7). Addition of calmodulin further enhanced the Ca2+ activating effect on 86Rb+ uptake (K+ channel activity). Ca2+-dependent 86Rb+ uptake was inhibited by tetraethylammonium ion and low pH. It is concluded that the adipocyte plasma membrane possesses K+ channels that are activated by Ca2+ and amplified by calmodulin.     

Electrophysiological responses to bradykinin and microinjected inositol polyphosphates in neuroblastoma cells. Possible role of inositol 1,3,4-trisphosphate in altering membrane potential

Addition of bradykinin to mouse N1E-115 neuroblastoma cells evokes a rapid but transient rise in cytoplasmic free Ca2+ concentration ([Ca2+]i). The [Ca2+]i rise is accompanied by a transient membrane hyperpolarization, due to a several-fold increase in K+ conductance, followed by a prolonged depolarizing phase. Pretreatment of the cells with a Ca2+-ionophore abolishes the hormone-induced hyperpolarization but leaves the depolarizing phase intact. The transient hyperpolarization can be mimicked by iontophoretic injection of IP3(1,4,5) or Ca2+, but not by injection of IP3(1,3,4), IP4(1,3,4,5) or Mg2+ into the cells. Instead, IP3(1,3,4) evokes a small but significant membrane depolarization in about 50% of the cells tested. Microinjected IP4(1,3,4,5) has no detectable effect, nor has treatment of the cells with phorbol esters. These results suggest that, while IP3(1,4,5) triggers the release of stored Ca2+ to hyperpolarize the membrane, IP3(1,3,4) may initiate a membrane depolarization.     

Inwardly rectifying current-voltage relationship of small-conductance Ca2+-activated K+ channels rendered by intracellular divalent cation blockade

Small conductance Ca2+-activated K+ channels (SK(Ca) channels) are a group of K+-selective ion channels activated by submicromolar concentrations of intracellular Ca2+ independent of membrane voltages. We expressed a cloned SK(Ca) channel, rSK2, in Xenopus oocytes and investigated the effects of intracellular divalent cations on the current-voltage (I-V) relationship of the channels. Both Mg2+ and Ca2+ reduced the rSK2 channel currents in voltage-dependent manners from the intracellular side and thus rectified the I-V relationship at physiological concentration ranges. The apparent affinity of Mg2+ was changed as a function of both transmembrane voltage and intracellular Ca2+ concentration. Extracellular K+ altered the voltage dependence as well as the apparent affinities of Mg2+ binding from intracellular side. Thus, the inwardly rectifying I-V relationship of SK(Ca) channels is likely due to the voltage-dependent blockade of intracellular divalent cations and that the binding site is located within the ion-conducting pathway. Therefore, intracellular Ca2+ modulates the permeation characteristics of SK(Ca) channels by altering the I-V relationship as well as activates the channel by interacting with the gating machinery, calmodulin, and SK(Ca) channels can be considered as Ca2+-activated inward rectifier K+ channels.     

Generation of action potentials in a mathematical model of corticotrophs.

Corticotropin-releasing hormone (CRH) is an important regulator of adrenocorticotropin (ACTH) secretion from pituitary corticotroph cells. The intracellular signaling system that underlies this process involves modulation of voltage-sensitive Ca2+ channel activity, which leads to the generation of Ca2+ action potentials and influx of Ca2+. However, the mechanisms by which Ca2+ channel activity is modulated in corticotrophs are not currently known. We investigated this process in a Hodgkin-Huxley-type mathematical model of corticotroph plasma membrane electrical responses. We found that an increase in the L-type Ca2+ current was sufficient to generate action potentials from a previously resting state of the model. The increase in the L-type current could be elicited by either a shift in the voltage dependence of the current toward more negative potentials, or by an increase in the conductance of the current. Although either of these mechanisms is potentially responsible for the generation of action potentials, previous experimental evidence favors the former mechanism, with the magnitude of the shift required being consistent with the experimental findings. The model also shows that the T-type Ca2+ current plays a role in setting the excitability of the plasma membrane, but does not appear to contribute in a dynamic manner to action potential generation. Inhibition of a K+ conductance that is active at rest also affects the excitability of the plasma membrane.     

Intracellular divalent cations and neuronal excitability.

Itracellular injections of Mg into cat spinal motoneurones have a depolarizing action, associated with a fall in input conductance, and depression of the postspike hyperpolarizing after-potential (a.h.p.) as well as its underlying conductance increase. There is also an increase in excitability, sometimes leading to outright discharge, and a change in the current-firing relation: the normal primary range is largely abolished and the firing appears to have the characteristics of the normal secondary range. Intracellular effects of Mg are thus mainly opposite to those of Ca, possibly owing to competition at sites where Ca activates K channels. Intracellular injections of Mn also tend to depress the a.h.p. but have relatively little effect on resting potential and conductance, or action potentials. Co also depresses the a.h.p. but has a more pronounced depolarizing action, and produces particularly strong depression of action potentials. By contrast intracellular Sr tends to raise the membrane conductance and has a mild hyperpolarizing effect. During the injection of Sr, a.h.p's are depressed but this is followed by a rebound of increased a.h.p. amplitude and conductance. Unlike the other divalent cations tested, Sr strongly depressed excitatory postsynaptic potentials. In most respects Sr appears to behave like Ca.     

Internal perfusion of guinea-pig hepatocytes with buffered Ca2+ or inositol 1,4,5-trisphosphate mimics noradrenaline activation of K+ and Cl- conductances

External application of noradrenaline to voltage-clamped guinea-pig isolated hepatocytes evoked membrane conductance increases to K+ and Cl-. This effect was reproduced by internal perfusion of the cells with 2 microM buffered Ca2+ and with 20 microM inositol 1,4,5-trisphosphate (IP3). The kinetic properties of the K+ conductance and its selective block by the toxin apamin were the same in each case. Cyclical fluctuations of conductance observed with noradrenaline were reproduced by internal IP3 but not by Ca2+ perfusion, indicating that oscillations of intracellular free Ca2+ may arise from properties of the Ca2+ sequestration mechanism at constant IP3 concentration.     

Photosensory transduction in the flagellated alga, Euglena gracilis I. Action of divalent cations, Ca2+ antagonists and Ca2+ ionophore on motility and photobehavior.

1. The flagellated alga, Euglena gracilis, swims forward essentially in a straight path under constant light intensity. Strong motility of the cells can be supported by Mg2+ alone but optimum motility is found in the presence of Mg2+, Ca2+ and K+. 2. Ca2+, Co2+, Mn2+ and Ba2+ induce a concentration-dependent increase in the rate at which the cells change the direction of their swimming path (a klinokinesis). Ni2+ immobilizes the flagellum. 3. On perception of a reduction ('step-down stimulus') in blue light intensity in their environment, Euglena rotate in place (tumble) for a finite period (the step-down photophobic response). 4. The duration of the tumbling is enhanced in the presence of divalent cations following the series Ca2+ greater than Ba2+ greater than Mn2+ greater than Co2+ greater than Mg2+ = Ni2+ = 0. 5. Neither the tumbling response in the presence of low concentrations of Ca2+ or the Ca2+-stimulated response is altered by verapamil (a Ca2+ conductance antagonist). The Ca2+ conductance/active transport antagonist, ruthenium red, is also inactive. 6. The Ca2+ ionophore, A23187, has little effect on flagellar activity in the absence of extracellular Ca2+. However, in the presence of A23187, Ca2+ induces a specific light-independent, concentration-dependent discontinuous tumbling response of the cells. 7. The data support a role for Ca2+ and Mg2+ in control of flagellar activity. However, blue light-induced tumbling behavior would not appear to be the direct result of a light-mediated alteration in the Ca2+ conductance of the flagellar membrane to affect flagellar reorientation. The results are discussed in connection with previous theories on control of flagella activity in green alga.     

Ionic mechanisms of histamine-induced responses in guinea pig intracardiac neurons

Histamine, released from mast cells, can modulate the activity of intrinsic neurons in the guinea pig cardiac plexus. The present study examined the ionic mechanisms underlying the histamine-induced responses in these cells. Histamine evokes a small membrane depolarization and an increase in neuronal excitability. Using intracellular voltage recording from individual intracardiac neurons, we were able to demonstrate that removal of extracellular sodium reduced the membrane depolarization, whereas inhibition of K+ channels by 1 mM Ba2+, 2 mM Cs+, or 5 mM tetraethylammonium had no effect. The depolarization was also not inhibited by either 10 microM Gd3+ or a reduced Cl- solution. The histamine-induced increase in excitability was unaffected by K+ channel inhibitors; however, it was reduced by either blockage of voltage-gated Ca2+ channels with 200 microM Cd2+ or replacement of extracellular Ca2+ with Mg2+. Conversely, alterations in intracellular calcium with thapsigargin or caffeine did not inhibit the histamine-induced effects. However, in cells treated with both thapsigargin and caffeine to deplete internal calcium stores, the histamine-induced increase in excitability was decreased. Treatment with the phospholipase C inhibitor U73122 also prevented both the depolarization and the increase in excitability. From these data, we conclude that histamine, via activation of H1 receptors, activates phospholipase C, which results in 1) the opening of a nonspecific cation channel, such as a transient receptor potential channel 4 or 5; and 2) in combination with either the influx of Ca2+ through voltage-gated channels or the release of internal calcium stores leads to an increase in excitability.     

Fatty acid modifies Ca2+-dependent potassium channel activity in smooth muscle cells from the human aorta

By using the patch-clamp technique the effect of 2-decenoic acid (DA) on Ca2+-activated potassium (K+) channels in the membrane of smooth muscle cells from the human aorta was studied. In the presence of 0.5 microM Ca2+ and 2 mM Mg2+ on the cytoplasmic side of the membrane, a more than tenfold elevation in the probability of the channels being open (po) was observed under the effect of DA. With divalent cation concentrations of less than 1 nM DA caused a more than twofold elevation in po. In the DA-treated membranes Mg2+ ions, which normally fail to activate the channels, brought about a nearly threefold increase in the channel activity when applied to the inner membrane surface. Channel sensitivity to the activating effect of cytoplasmic Ca2+ ions did not increase with the application of DA. Single-channel conductance was unchanged by DA exposure. We suggest that DA alters the Ca2+-binding mechanism of the channel, increasing its sensitivity to Mg2+ ions, presumably owing to membrane fluidization.     

Role of exercise-induced potassium fluxes underlying muscle fatigue: a brief review

The site of exercise-induced muscle fatigue is suggested to be the muscle membrane, which includes the sarcolemma and T-tubule membrane; the excitability of the membrane is dependent on the membrane potential. Significant potassium flux from the intracellular space of contracting muscle may decrease the membrane potential to half its resting value. This is true for isolated muscle preparations as well as for the whole body exercise in humans. Specific K+ channels have been identified, that may account for the intracellular K+ loss. Calcium-sensitive K+ channels open when intracellular Ca2+ concentrations increase, as during excitation. ATP-sensitive K+ channels may be involved but may open only at ATP concentrations well below those attained at exhaustion. However, ATP may be compartmentalized and only the membrane-bound ATP concentration may be of significance. Ca2+ accumulation and ATP depletion cause cell destruction; these changes induce an increased K+ conductance, which may inactivate the membrane and consequently prevent tension development. It is hypothesized that such a safety mechanism is identical to the fatigue mechanism.     

BKCa channels activating at resting potential without calcium in LNCaP prostate cancer cells

Large-conductance Ca2+-dependent K+ (BK(Ca)) channels are activated by intracellular Ca2+ and membrane depolarization in an allosteric manner. We investigated the pharmacological and biophysical characteristics of a BK(Ca)-type K+ channel in androgen-dependent LNCaP (lymph node carcinoma of the prostate) cells with novel functional properties, here termed BK(L). K+ selectivity, high conductance, activation by Mg2+ or NS1619, and inhibition by paxilline and penitrem A largely resembled the properties of recombinant BK(Ca) channels. However, unlike conventional BK(Ca) channels, BK(L) channels activated in the absence of free cytosolic Ca2+ at physiological membrane potentials; the half-maximal activation voltage was shifted by about -100 mV compared with BK(Ca) channels. Half-maximal Ca2+-dependent activation was observed at 0.4 microM: for BK(L) (at -20 mV) and at 4.1 microM: for BK(Ca) channels (at +50 mV). Heterologous expression of hSlo1 in LNCaP cells increased the BK(L) conductance. Expression of hSlo-beta1 in LNCaP cells shifted voltage-dependent activation to values between that of BK(L) and BK(Ca) channels and reduced the slope of the P (open) (open probability)-voltage curve. We propose that LNCaP cells harbor a so far unknown type of BK(Ca) subunit, which is responsible for the BK(L) phenotype in a dominant manner. BK(L)-like channels are also expressed in the human breast cancer cell line T47D. In addition, functional expression of BK(L) in LNCaP cells is regulated by serum-derived factors, however not by androgens.     

Electrical pacemaker mechanisms of pancreatic islet cells

Glucose, the major physiological stimulus for insulin secretion, induces a periodic bursting pattern of Ca2+ action potentials that are thought to mediate the uptake of Ca2+ into the intracellular pool of free Ca2+, which controls the rate of insulin release. Evidence is reviewed that shows that the voltage-dependent Ca2+ spikes are driven by a slow, voltage-dependent plateau depolarization that may also be caused by Ca2+ influx. Current evidence suggests that this plateau conductance is periodically terminated in turn by a pacemaker current through membrane K+ channels that are activated by intracellular free Ca2+. The control of electrical activity by different modulators of insulin release may involve interactions with this system at several points, including changes of the sensitivity of K+ channels to intracellular Ca2+ and to changes of intracellular Ca2+ buffering capacity.     

A high-affinity, calmodulin-sensitive (Ca2+ + Mg2+)-ATPase and associated calcium-transport pump in the Ehrlich ascites tumor cell plasma membrane

A unique cytoplast preparation from Ehrlich ascites tumor cells (G. V. Henius, P. C. Laris, and J. D. Woodburn (1979) Exp. Cell. Res. 121, 337-345), highly enriched in plasma membranes, was employed to characterize the high-affinity plasma membrane calcium-extrusion pump and its associated adenosine triphosphatase (ATPase). An ATP-dependent calcium-transport system which had a high affinity for free calcium (K0.5 = 0.040 +/- 0.005 microM) was identified. Two different calcium-stimulated ATPase activities were detected. One had a low (K0.5 = 136 +/- 10 microM) and the other a high (K0.5 = 0.103 +/- 0.077 microM) affinity for free calcium. The high-affinity enzyme appeared to represent the ubiquitous high-affinity plasma membrane (Ca2+ + Mg2+)-ATPase (calcium-stimulated, magnesium-dependent ATPase) seen in normal cells. Both calcium transport and the (Ca2+ + Mg2+)-ATPase were significantly stimulated by the calcium-dependent regulatory protein calmodulin, especially when endogenous activator was removed by treatment with the calcium chelator ethylene glycol bis(beta-aminoethyl ether) N,N'-tetraacetic acid. Other similarities between calcium transport and the (Ca2+ + Mg2+)-ATPase included an insensitivity to ouabain (0.5 mM), lack of activation by potassium (20 mM), and a requirement for magnesium. These similar properties suggested that the (Ca2+ + Mg2+)-ATPase represents the enzymatic basis of the high-affinity calcium pump. The calcium pump/enzyme system was inhibited by orthovanadate at comparatively high concentrations (calcium transport: K0.5 congruent to 100 microM; (Ca2+ + Mg2+)-ATPase: K0.5 greater than 100 microM). Upon Hill analysis, the tumor cell (Ca2+ + Mg2+)-ATPase failed to exhibit cooperative activation by calcium which is characteristic of the analogous enzyme in the plasma membrane of normal cells.     

A lineage-specific Ca(2+)-activated K+ conductance in HL-60 cells.

Cells of the human promyelocytic cell line HL-60 can be controllably induced to terminally differentiate into either granulocytes or monocyte/macrophages. HL-60 promyelocytes and terminally differentiated macrophages express a K(+)-selective ion channel which is activated by intracellular free Ca2+ concentrations above 10(-7) M. Because of its voltage independence, this channel can be distinguished from the voltage- and Ca(2+)-activated family of outward-rectifying channels. The channel is selective for K+ against Na+ and is blocked by Ba2+, thus it may be similar to the Ca(2+)-activated K+ channel previously described in human macrophages. In its sensitivity to block by charybdotoxin, this channel also resembles a Ca(2+)-activated K+ channel of lymphocytes, which plays a role in activation-dependent hyperpolarization. In contrast to promyelocytes and macrophages, functional expression of the Ca(2+)-activated K+ channel is suppressed to nearly undetectable levels in granulocytes derived from HL-60 cells by retinoic acid-induced differentiation. These data suggest that signals which produce elevation of intracellular Ca2+ will hyperpolarize promyelocytes and differentiated macrophages by activating this conductance; however, signals which elevate free Ca2+ in granulocytes must act on other effectors, which may produce a different final influence on membrane potential.     

Ionic regulation of adenylate cyclase from the cilia of Paramecium tetraurelia.

The kinetics of the ionic regulation of an adenylate cyclase associated with the excitable ciliary membrane from Paramecium tetraurelia was examined. Glycerol (30%, v/v) stabilized the enzyme, and activated by an increase in Vmax. (3-fold) and a decrease in the apparent Km for MgATP (6-fold). Kinetic analysis of Mg2+ effects showed a stimulation via a single metal-binding site separate from the substrate site, with a dissociation constant, Ks, of 0.27 mM. Analysis of Ca2+ effects showed (i) an uncompetitive inhibition with respect to substrate MgATP, and (ii) dependence of the extent of inhibition on the free Mg2+ concentration. Ki values ranged from 4 to 130 microM-Ca2+ in the presence of 0.55-2 mM-Mg2+ respectively. This indicates competition between Mg2+ and Ca2+ at the metal-binding site. The Ca2+ effect was specific; Sr2+ and Ba2+ were almost without effect, and 100 microM-Ba2+ did not interfere with the Ca2+ inhibition. The actions of Ca2+ were readily reversible after addition of EGTA. K+ activated the adenylate cyclase at concentrations around 20 mM. The stimulatory potency of K+ was dependent on the free Mg2+ concentration. At 1 mM free Mg2+, 20 mM-K+ doubled the adenylate cyclase activity. The inhibitory Ca2+ and stimulatory K+ inputs were independent of each other.     

Magnesium permeability of sarcoplasmic reticulum. Mg2+ is not countertransported during ATP-dependent Ca2+ uptake by sarcoplasmic reticulum

Magnesium transport across sarcoplasmic reticulum (SR) vesicles was investigated in reaction mixtures of various composition using antipyrylazo III or arsenazo I to monitor extravesicular free Mg2+. The half-time of passive Mg2+ efflux from Mg2+-loaded SR was 100 s in 100 mM KCl, 150 S in 100 mM K gluconate, and 370 S in either 100 mM Tris methanesulfonate or 200 mM sucrose solutions. The concentration and time course of Mg2+ released into the medium was also measured during ATP-dependent Ca2+ uptake by SR. In reaction mixtures containing up to 3 mM Mg2+, small changes in free magnesium of 10 microM or less were accurately detected without interference from changes in free Ca2+ of up to 100 microM. Three experimental protocols were used to determine whether the increase of free [Mg2+] in the medium after an addition of ATP was due to Mg2+ dissociated from ATP following ATP hydrolysis or to Mg2+ translocation from inside to outside of the vesicles. 1) In the presence of ATP-regenerating systems which maintained constant ATP to ADP ratios and normal rates of active Ca2+ uptake, the increase of Mg2+ in the medium was negligible. 2) Mg2+ released during ATP-dependent Ca2+ uptake by SR was similar to that observed during ATP hydrolysis catalyzed by apyrase, in the absence of SR. 3) In SR lysed with Triton X-100 such that Ca2+ transport was uncoupled from ATPase activity, the rate and amount of Mg2+ release was greater than that observed during ATP-dependent Ca2+ uptake by intact vesicles. Taken together, the results indicate that passive fluxes of Mg2+ across SR membranes are 10 times faster than those of Ca2+ and that Mg2+ is not counter-transported during active Ca2+ accumulation by SR even in reaction mixtures containing minimal concentrations of membrane permeable ions that could be rapidly exchanged or cotransported with Ca2+ (e.g. K+ or Cl-).