Cation-selective channels in amphibian epithelia: electrophysiological properties and activation

1. Na+ as well as Li+ move across the apical membrane through amiloride-sensitive ionic channels. 2. K+ movements across the apical membrane occur through Ba2+- and Cs+-sensitive channels which do not allow the passage of Na+ or Li+. 3. A third pathway in the apical membrane is permeable for Na+, K+, Cs+, Rb+, NH+4 and Ti+. The currents carried by these monovalent cations are blocked by Ca2+ and divalent cations as well as La3+. 4. In the urinary bladder, the Ca2+-sensitive currents are stimulated by oxytocin, activators of cytosolic cAMP and cAMP analogues. Also the oxytocin activated currents are blocked by divalent cations and La3+. 5. Nanomolar concentrations of mucosal Ag+ activate the third channel and open the pathway for movements of Ca2+, Ba2+ and Mg2+, which are known to permeate through Ca2+ channels in excitable tissues.     

Na+/Ca2+ antiport in cultured arterial smooth muscle cells. Inhibition by magnesium and other divalent cations

Cultured smooth muscle cells from rat aorta were loaded with Na+, and Na+/Ca2+ antiport was assayed by measuring the initial rates of 45Ca2+ influx and 22Na+ efflux, which were inhibitable by 2',4'-dimethylbenzamil. The replacement of extracellular Na+ with other monovalent ions (K+, Li+, choline, or N-methyl-D-glucamine) was essential for obtaining significant antiport activity. Mg2+ competitively inhibited 45Ca2+ influx via the antiporter (Ki = 93 +/- 7 microM). External Ca2+ or Sr2+ stimulated 22Na+ efflux as would be expected for antiport activity. Mg2+ did not stimulate 22Na+ efflux, which indicates that Mg2+ is probably not transported by the antiporter under the conditions of these experiments. Mg2+ inhibited Ca2+-stimulated 22Na+ efflux as expected from the 45Ca2+ influx data. The replacement of external N-methyl-D-glucamine with K+, but not other monovalent ions (choline, Li+), decreased the potency of Mg2+ as an inhibitor of Na+/Ca2+ antiport 6.7-fold. Other divalent cations (Co2+, Mn2+, Cd2+, Ba2+) also inhibited Na+/Ca2+ antiport activity, and high external potassium decreased the potency of each by 4.3-8.6-fold. The order of effectiveness of the divalent cations as inhibitors of Na+/Ca2+ antiport (Cd2+ greater than Mn2+ greater than Co2+ greater than Ba2+ greater than Mg2+) correlated with the closeness of the crystal ionic radius to that of Ca2+.     

Sodium permeability of a cloned small-conductance calcium-activated potassium channel

Small-conductance Ca2+-activated potassium channels (SK(Ca) channels) are heteromeric complexes of pore-forming main subunits and constitutively bound calmodulin. SK(Ca) channels in neuronal cells are activated by intracellular Ca2+ that increases during action potentials, and their ionic currents have been considered to underlie neuronal afterhyperpolarization. However, the ion selectivity of neuronal SK(Ca) channels has not been rigorously investigated. In this study, we determined the monovalent cation selectivity of a cloned rat SK(Ca) channel, rSK2, using heterologous expression and electrophysiological measurements. When extracellular K+ was replaced isotonically with Na+, ionic currents through rSK2 reversed at significantly more depolarized membrane potentials than the value expected for a Nernstian relationship for K+. We then determined the relative permeability of rSK2 for monovalent cations and compared them with those of the intermediate- and large-conductance Ca2+-activated K+ channels, IK(Ca) and BK(Ca) channels. The relative permeability of the rSK2 channel was determined as K+(1.0)>Rb+(0.80)>NH(4)+(0.19) approximately Cs+(0.19)>Li+(0.14)>Na+(0.12), indicating substantial permeability of small ions through the channel. Although a mutation near the selectivity filter mimicking other K+-selective channels influenced the size-selectivity for permeant ions, Na+ permeability of rSK2 channels was still retained. Since the reversal potential of endogenous SK(Ca) current is determined by Na+ permeability in a physiological ionic environment, the ion selectivity of native SK(Ca) channels should be reinvestigated and their in vivo roles may need to be restated.     

Conductance and permeation of monovalent cations through depletion-activated Ca2+ channels (ICRAC) in Jurkat T cells.

We studied monovalent permeability of Ca2+ release-activated Ca2+ channels (ICRAC) in Jurkat T lymphocytes following depletion of calcium stores. When external free Ca2+ ([Ca2+]o) was reduced to micromolar levels in the absence of Mg2+, the inward current transiently decreased and then increased approximately sixfold, accompanied by visibly enhanced current noise. The monovalent currents showed a characteristically slow deactivation (tau = 3.8 and 21.6 s). The extent of Na+ current deactivation correlated with the instantaneous Ca2+ current upon readdition of [Ca2+]o. No conductance increase was seen when [Ca2+]o was reduced before activation of ICRAC. With Na+ outside and Cs+ inside, the current rectified inwardly without apparent reversal below 40 mV. The sequence of conductance determined from the inward current at -80 mV was Na+ > Li+ = K+ > Rb+ >> Cs+. Unitary inward conductance of the Na+ current was 2.6 pS, estimated from the ratios delta sigma2/delta Imean at different voltages. External Ca2+ blocked the Na+ current reversibly with an IC50 value of 4 microM. Na+ currents were also blocked by 3 mM Mg2+ or 10 microM La3+. We conclude that ICRAC channels become permeable to monovalent cations at low levels of external divalent ions. In contrast to voltage-activated Ca2+ channels, the monovalent conductance is highly selective for Na+ over Cs+. Na+ currents through ICRAC channels provide a means to study channel characteristics in an amplified current model.     

Regulation of connexin hemichannels by monovalent cations

Opening of connexin hemichannels in the plasma membrane is highly regulated. Generally, depolarization and reduced extracellular Ca2+ promote hemichannel opening. Here we show that hemichannels formed of Cx50, a principal lens connexin, exhibit a novel form of regulation characterized by extraordinary sensitivity to extracellular monovalent cations. Replacement of extracellular Na+ with K+, while maintaining extracellular Ca2+ constant, resulted in >10-fold potentiation of Cx50 hemichannel currents, which reversed upon returning to Na+. External Cs+, Rb+, NH4+, but not Li+, choline, or TEA, exhibited a similar effect. The magnitude of potentiation of Cx50 hemichannel currents depended on the concentration of extracellular Ca2+, progressively decreasing as external Ca2+ was reduced. The primary effect of K+ appears to be a reduction in the ability of Ca2+, as well as other divalent cations, to close Cx50 hemichannels. Cx46 hemichannels exhibited a modest increase upon substituting Na+ with K+. Analyses of reciprocal chimeric hemichannels that swap NH2- and COOH-terminal halves of Cx46 and Cx50 demonstrate that the difference in regulation by monovalent ions in these connexins resides in the NH2-terminal half. Connexin hemichannels have been implicated in physiological roles, e.g., release of ATP and NAD+ and in pathological roles, e.g., cell death through loss or entry of ions and signaling molecules. Our results demonstrate a new, robust means of regulating hemichannels through a combination of extracellular monovalent and divalent cations, principally Na+, K+, and Ca2+.     

Activation of calcium transport in skeletal muscle sarcoplasmic reticulum by monovalent cations.

The rates of calcium transport and Ca2+-dependent ATP hydrolysis by rabbit skeletal muscle sarcoplasmic reticulum were stimulated by monovalent cations. The rate of decomposition of phosphoprotein intermediate of the Ca2+-dependent ATPase of sarcoplasmic reticulum was also increased by these ions to an extent that is sufficient to account for the stimulation of calcium transport and Ca2+-dependent ATPase activity. The order of effectiveness of monovalent cations tested at saturating concentrations in increasing rate of phosphoprotein decomposition is: K+, Na+ greater than Rb+, NH4+ greater than Cs+ greater than Li+, choline+, Tris+.     

Permeation properties of a Ca(2+)-blockable monovalent cation channel in the ectoderm of the chick embryo: pore size and multioccupancy probed with organic cations and Ca2+

A Ca(2+)-blockable monovalent cation channel is present in the apical membrane of the ectoderm of the gastrulating chick embryo. We used the patch clamp technique to study several single-channel permeation properties of this channel. In symmetrical conditions without Ca2+, the Na+ current carried by the channel rectifies inwardly. The channel has an apparent dissociation constant for extracellular Na+ of 115 mM at 0 mV and a low density of negative surface charge (-0.03 e/nm2) at its extracellular entrance. The minimal pore diameter is approximately 5.8 A, as calculated from the relative permeabilities of 10 small organic cations. Extracellular application of six large organic cations decreased the inward Na+ current in a voltage-dependent manner, which strongly suggests an intrachannel block. The presence of at least two ion binding sites inside the pore is inferred from the Na+ dependence of the block by the organic cations. This hypothesis is strengthened by the fact that the extracellular Ca2+ block is also modified by the Na+ concentration. In particular, the rise of the unblocking rate with increased Na+ concentrations clearly suggests the presence of an interaction between Ca2+ and Na+ inside the channel. A low probability of double occupancy at physiological ionic conditions is implied from the absence of an anomalous mole fraction effect with mixtures of extracellular Li+ and K+. Finally, the absence of inward current at very strong hyperpolarizations and in the presence of 10 mM extracellular Ca2+ demonstrates the absence of significant Ca2+ current through this channel. It is argued that this embryonic epithelial Ca(2+)-blockable monovalent cation channel is related to both L-type Ca2+ channel and cyclic nucleotide-gated channels.     

Potassium current inhibition by nonselective cation channel-mediated sodium entry in rat pheochromocytoma (PC-12) cells.

Under physiological conditions, nonselective cation (NSC) channels mediate the entry of cations into cells, the most important being Na+ and Ca2+. In contrast to the Ca(2+)-dependent signaling mechanisms, little is known about the consequences and the spatial distribution of intracellular [Na+] elevation. In this study we demonstrate that Na+ entry, during the opening of ATP-activated NSC channels, leads to an inhibition of voltage-dependent K+ currents (IK) in cromaffin-like undifferentiated PC-12 cells. The effect was dependent on the charge carrier as well as on the density of the ATP-activated current. Extracellular alkali cations (Na+, Li+) were more efficient than NH4+ in suppressing IK. Intracellular infusion of Na+ had the same effect as Na+ influx through ATP-activated NSC channels. The inhibition of IK persisted when the total ATP-induced Na+ entry was reduced by membrane depolarization, suggesting a spatial restriction of the required Na+ accumulation. Our results indicate that NSC channels influence the function of other ion channels by changing local intracellular ion concentrations.     

Cationic modulation of follicle-stimulating hormone binding to granulosa cell receptor

Magnesium (Mg2+) increases binding of follicle-stimulating hormone (FSH) to membrane-bound receptors and increases adenylyl cyclase activity. We examined the effects of divalent and monovalent cations on FSH binding to receptors in granulosa cells from immature porcine follicles. Divalent and monovalent cations increased binding of [125I]iodo-porcine FSH (125I-pFSH). The divalent cations Mg2+, calcium (Ca2+) and manganese, (Mn2+) increased specific binding a maximum of 4- to 5-fold at added concentrations of 10 mM. Mg2+ caused a half-maximal enhancement of binding at 0.6 mM, whereas Ca2+ and Mn2+ had half-maximal effects at 0.7 mM and 0.8 mM, respectively. The monovalent cation potassium (K+) increased binding a maximum of 1.5-fold at an added concentration of 50 mM, whereas the monovalent cation (Na+) did not increase binding at any concentration tested. The difference between K+ and Na+ suggested that either enhancement of binding was not a simple ionic effect or Na+ has a negative effect that suppresses its positive effect. Ethylenediamine tetraacetic acid, a chelator of Mg2+, prevented binding of 125I-pFSH only in the presence of Mg2+, whereas pregnant mare's serum gonadotropin, a competitor with FSH for the receptor, prevented binding in both the absence and the presence of Mg2+. Guanyl-5-ylimidodiphosphate (Gpp[NH]p) inhibited binding of 125I-pFSH in the absence or presence of Mg2+, but only at Gpp(NH)p concentrations greater than 1 mM. We used Mg2+ to determine if divalent cations enhanced FSH binding by increasing receptor affinity or by increasing the apparent number of binding sites.(ABSTRACT TRUNCATED AT 250 WORDS)     

Gill (Na+ + K+)- and Na+-stimulated Mg2+-dependent ATPase activities in the gilthead bream (Sparus auratus L.)

1. Gilthead gill 10(-3) M ouabain-inhibited (Na+ + K+)-ATPase and 10(-2) M ouabain-insensitive Na+-ATPase require the optimal conditions of pH 7.0, 160 mM Na+, 20 mM K+, 5 mM MgATP and pH 4.8-5.2, 75 mM Na+, 2.5 mM Mg2+, 1.0 mM ATP, respectively. 2. The main distinctive features between the two activities are confirmed to be optimal pH, the ouabain-sensitivity and the monovalent cation requirement, Na+ plus another cationic species (K+, Rb+, Cs+, NH4+) in the (Na+ + K+)-ATPase and only one species (Na+, K+, Li+, Rb+, Cs+, NH4+ or choline+) in the Na+-ATPase. 3. The aspecific Na+-ATPase activation by monovalent cations, as well as by nucleotide triphosphates, opposed to the (Na+ + K+)-ATPase specificity for ATP and Na+, relates gilthead gill ATPases to lower organism ATPases and differentiates them from mammalian ones. 4. The discrimination between the two activities by the sensitivity to ethacrynic acid, vanadate, furosemide and Ca2+ only partially agrees with the literature. 5. Present findings are viewed on the basis of the ATPase's presumptive physiological role(s) and mutual relationship.     

Effect of Monovalent Cations on Na+/Ca2+ Exchange and ATP-Dependent Ca2+ Transport in Synaptic Plasma Membranes

Two Ca2+ transport systems were investigated in plasma membrane vesicles isolated from sheep brain cortex synaptosomes by hypotonic lysis and partial purification. Synaptic plasma membrane vesicles loaded with Na+ (Na+i) accumulate Ca2+ in exchange for Na+, provided that a Na+ gradient (in leads to out) is present. Agents that dissipate the Na+ gradient (monensin) prevent the Na+/Ca2+ exchange completely. Ca2+ accumulated by Na+/Ca2+ exchange can be released by A 23187, indicating that Ca2+ is accumulated intravesicularly. In the absence of any Na+ gradient (K+i-loaded vesicles), the membrane vesicles also accumulate Ca2+ owing to ATP hydrolysis. Monovalent cations stimulate Na+/Ca2+ exchange as well as the ATP-dependent Ca2+ uptake activity. Taking the value for Na+/Ca2+ exchange in the presence of choline chloride (external cation) as reference, other monovalent cations in the external media have the following effects: K+ or NH4+ stimulates Na+/Ca2+ exchange; Li+ or Cs+ inhibits Na+/Ca2+ exchange. The ATP-dependent Ca2+ transport system is stimulated by increasing K+ concentrations in the external medium (Km for K+ is 15 mM). Replacing K+ by Na+ in the external medium inhibits the ATP-dependent Ca2+ uptake, and this effect is due more to the reduction of K+ than to the elevation of Na+. The results suggest that synaptic membrane vesicles isolated from sheep brain cortex synaptosomes possess mechanisms for Na+/Ca2+ exchange and ATP-dependent Ca2+ uptake, whose activity may be regulated by monovalent cations, specifically K+, at physiological concentrations.     

Interaction of sodium and potassium ions with Na+,K+-ATPase. II. General properties of ouabain-sensitive K+ binding

General properties of ouabain-sensitive K+ binding to purified Na+,K+-ATPase [EC 3.6.1.3] were studied by a centrifugation method with 42K+. 1) The affinity for K+ was constant at pH values higher than 6.4, and decreased at pH values lower than 6.4. 2) Mg2+ competitively inhibited the K+ binding. The dissociation constant (Kd) for Mg2+ of the enzyme was estimated to be about 1 mM, and the ratio of Kd for Mg2+ to Kd for K+ was 120 : 1. The order of inhibitory efficiency of divalent cations toward the K+ binding was Ba2+ congruent to Ca2+ greater than Zn2+ congruent to Mn2+ greater than Sr2+ greater than Co2+ greater than Ni2+ greater than Mg2+. 3) The order of displacement efficiency of monovalent cations toward the K+ binding in the presence or absence of Mg2+ was Tl+ greater than Rb+ greater than or equal to (K+) greater than NH4+ greater than or equal to Cs+ greater than Na+ greater than Li+. The inhibition patterns of Na+ and Li+ were different from those of other monovalent cations, which competitively inhibited the K+ binding. 4) The K+ binding was not influenced by different anions, such as Cl-, SO4(2-), NO3-, acetate, and glycylglycine, which were used for preparing imidazole buffers. 5) Gramicidin D and valinomycin did not affect the K+ binding, though the former (10 micrograms/ml) inhibited the Na+,K+-ATPase activity by about half. Among various inhibitors of the ATPase, 0.1 mM p-chloromercuribenzoate and 0.1 mM tri-n-butyltin chloride completely inhibited the K+ binding. Oligomycin (10 micrograms/ml) and 10 mM N-ethylmaleimide had no effect on the K+ binding. In the presence of Na+, however, oligomycin decreased the K+ binding by increasing the inhibitory effect of Na+, whether Mg2+ was present or not. 6) ATP, adenylylimido diphosphate and ADP each at 0.2 mM decreased the K+ binding to about one-fourth of the original level at 10 microM K+ without MgCl2 and at 60 microM K+ with 5 mM MgCl2. On the other hand, AMP, Pi, and p-nitrophenylphosphate each at 0.2 mM had little effect on the K+ binding.     

Interactions of metal ions with phosphatidylserine bilayer membranes: effect of hydrocarbon chain unsaturation

A combination of surface monolayer, scanning calorimetry, 31P NMR, and spin-label ESR techniques has been used to monitor the interactions of monovalent (NH4+, Na+, and Li+) and divalent (Ca2+) cations with phosphatidylserines (PS) differing in their levels of chain unsaturation. Comparisons are made between the disaturated dimyristoyl-, dipalmitoyl-, and dihexadecyl-PS (DMPS, DPPS, and DHPS), saturated cis-monounsaturated palmitoyloleoyl-PS (POPS) (and bovine brain PS), di-trans-monounsaturated dielaidoyl-PS (DEPS), and di-cis-monounsaturated dioleoyl-PS (DOPS). Na+ and NH4+ cations interact weakly with all PS monolayers and bilayers without significant changes in molecular conformation, chain packing, or headgroup dynamics and without dependence on chain composition. In contrast, considering these structural and dynamic parameters, Li+ shows a gradation in its interaction with PS (DMPS greater than POPS approximately bovine brain PS greater than DOPS), suggesting that Li+-PS interactions depend on the interfacial properties of the PS molecules (e.g., surface area). Finally, Ca2+ interacts strongly with all PS monolayers and bilayers, without obvious chain selectivity. Thus, ion binding to PS depends not only on the properties of the cation (Na+ vs Li+ vs Ca2+) but also on the molecular details of the PS membrane surface.     

Dependence of Na+-Ca2+ exchange and Ca2+-Ca2+ exchange on monovalent cations

Two mechanisms of passive Ca2+ transport, Na+-Ca2+ exchange and Ca2+-Ca2+ exchange, were studied using highly-purified dog heart sarcolemmal vesicles. About 80% of the Ca2+ accumulated by Na+-Ca2+ exchange or Ca2+-Ca2+ exchange could be released as free Ca2+, while up to 20% was probably bound. Na+-Ca2+ exchange was simultaneous, coupled countertransport of Na+ and Ca2+. The movement of anions during Na+-Ca2+ exchange did not limit the initial rate of Na+-Ca2+ exchange. Na+-Ca2+ exchange was electrogenic, with a reversal potential of about -105 mV. The apparent flux ratio of Na+-Ca2+ exchange was 4 Na+:1 Ca2+. Coupled cation countertransport by the Na+-Ca2+ exchange mechanism required a monovalent cation gradient with the following sequence of ion activation: Na+ much greater than Li+ greater than Cs+ greater than K+ greater than Rb+. In contrast to Na+-Ca2+ exchange, Ca2+-Ca2+ exchange did not require a monovalent cation gradient, but required the presence of Ca2+ plus a monovalent cation on both sides of the vesicle membrane. The sequence of ion activation of Ca2+-Ca2+ exchange was: K+ much greater than Rb+ greater than Na+ greater than Li+ greater than Cs+. Na+ inhibited Ca2+-Ca2+ exchange when Ca2+-Ca2+ exchange was supported by another monovalent cation. Both Na+-Ca2+ exchange and Ca2+-Ca2+ exchange were inhibited, but with different sensitivities, by external MgCl2, quinidine, or verapamil.     

Cyclic nucleotide-activated channels in the frog olfactory receptor plasma membrane.

Patch clamp technique was used to record cyclic nucleotide-dependent current of the frog olfactory receptor cell plasma membrane. Data obtained indicate that the channels passing this current are permeable to Ca2+ or Mg2+ and moderately selective for monovalent cations according to the sequence Li+, Na+, K+ greater than Rb+ greater than Cs+ and are effectively blocked by 1-cis-diltiazem and 3',4'-dichlorobenzamil. The conductance of single cyclic nucleotide-gated channels in solutions with low Ca2+ and Mg2+ content is about 19 pS. The results demonstrate that cyclic nucleotide-activated channels of olfactory receptor cells are virtually identical to photoreceptor ones.     

The effects of storage of sarcoplasmic reticulum fragments on the Ca2+, Mg2+-ATPase.

The effects of K+ and Na+ on the Ca2+,Mg2+-ATPase of sarcoplasmic reticulum fragments (SRF) were investigated at 1 mM ATP. There was an alteration of the sensitivity of the ATPase to the monovalent cations during storage of the SRF preparation. The Ca2+, Mg2+-ATPase of freshly prepared SRF was slightly activated by 5-10 mM K+ and Na+. Mg2+-ATPase was inhibited by both the monovalent cations to the same extent, and this response to the ions was independent of the freshness of the preparations. After storage of SRF, however, the Ca2+,Mg2+-ATPase was markedly activated by higher concentrations of K+ and Na+ (0.2-0.3 M). K+ and Na+ reduced the Ca uptake at the steady state in freshly prepared SRF, but did not affect pre-steady state uptake. In the presence of oxalate, the rate of Ca accumulation both in fresh and stored preparations was activated by 0.1-0.2 M K+ and Na+. The Ca2+, mg2+-ATPase with oxalate, so-called "extra ATPase," showed the same response to the ions as did the activity without oxalate during storage.     

Condensation of the chromatin gel by cations

Calf thymus chromatin gel, containing strongly bound nonhistone proteins, was used to study the effect of easily removable and tightly bound cations on the condensation of chromatin. The chromatin volume was found to be linearly dependent on the reciprocal square root of the concentration of easily removable cations (Tris X H+ + Na+ and Mg2+) except for the initial stages of condensation (up to 7-10 mM monovalent and 0.15-0.2 mM divalent cations). The effect of Mg2+ at the initial stage of condensation was not reproduced by Na+ and vice versa. At higher concentrations the effects of Na+ and Mg2+ were additive. The removal of tightly bound divalent cations by a treatment of the chromatin gel with 1,10-phenanthroline led to an approx. 50% increase in the volume of the chromatin gel, which was maintained at each concentration of easily removable cations. The 1,10-phenanthroline-caused decondensation of the chromatin gel was reversed by Ca2+ but not by Mg2+, Zn2+ and Cu2+. The chromatin gel pretreated with Ca2+ was not further decondensed by 1,10-phenanthroline.     

Cu2+-induced permeability of the Escherichia coli cytoplasmic membrane]

Cu(2+)-induced permeability of cytoplasmic membranes of Escherichia coli for different cations and neutral molecules of saccharose was estimated by studying their effect on cell plasmolysis during uncharged exchange of cytoplasmic K+ ions by periplasmic space cations. The addition of copper resulted in the exchange of K+ ions by periplasmic Na+, Tris+, streptomycin2+, Cu2+, Ca2+, Mg2+, Cd2+, and Mn2+. It is concluded that Cu(2+)-induced conducting pathways in bacterial membranes are hydrophilic channels with a radius of approximately 0.5 nm and a nonselective permeability for different cations.     

Ca(2+)-blockable, poorly selective cation channels in the apical membrane of amphibian epithelia. UO2(2+) reveals two channel types

This study deals with the effect of mucosal UO2(2+) on the Ca(2+)- blockable, poorly selective cation channels in the apical membrane of frog skin and toad urinary bladder. Our data show that UO2(2+) inhibits the Na+ currents through the amiloride-insensitive cation pathway and confirm a previously described stimulatory effect on the amiloride- blockade Na+ transport. Noise analysis of the Ca(2+)-blockable current demonstrates that the divalent also depresses the low-frequency Lorentzian (fc = 11.7 Hz) in the power density spectrum (PDS) and reveals the presence of high-frequency relaxation noise (fc = 58.5 Hz). The action of UO2(2+) is not reversed upon washout and is not accompanied by noise, typically induced by reversible blockers. The divalent merely depresses the plateau of the low-frequency Lorentzian, demonstrating a decrease in the number of conductive cation channels. Similarly, with mucosal K+ and Rb+, UO2(2+) also unmasks the high- frequency Lorentzian by depressing the noise from the slowly fluctuating cation channels (type S). In all experiments with mucosal Cs+, the PDS contains high-frequency relaxation noise (fc = 75.1 Hz in Rana temporaria, and 65.4 Hz in Rana ridibunda). An effect of UO2(2+) on the Cs+ currents and Lorentzian plateaus could not be demonstrated, suggesting that this monovalent cation does not pass through type S channels. Experiments with the urinary bladder revealed only a UO2(2+)- insensitive pathway permeable for Na+, K+, Rb+, and Cs+. We submit that in frog skin two cation-selective channels occur, distinguished by their spontaneous gating kinetics, their sensitivity to UO2(2+), and their permeability for Cs+. In toad urinary bladder, only one kind of cation-selective channel is observed, which resembles the UO2(2+)- insensitive channel in frog skin, with fast open-closed kinetics (type F).     

Ionic currents of channels that are permeable to monovalent and divalent cations.

The cation-selective channel from Tetrahymena cilia is permeable to both monovalent and divalent cations. The single channel conductance in mixed solutions of K+ and Ca2+ was determined by the Gibbs-Donnan ratio of K+ and Ca2+, and the binding sites of this channel were considered to be always occupied by two potassium ions or by one calcium ion under the experimental conditions: 5-90 mM K+ and 0.5-35 mM Ca2+ (Oosawa and Kasai, 1988). A two-barrier model for the channel was introduced and the values of Michaelis-Menten constants and maximum currents carried by K+ and Ca2+ were calculated using this model. Single channel current amplitudes and reversal potentials were calculated from these values. The calculated single-channel currents were compared with those obtained experimentally. The calculated reversal potentials were compared with the resting potentials of Tetrahymena measured in various concentrations of extracellular K+ and Ca2+. The method of calculation of ionic currents and reversal potentials presented here is helpful for understanding the properties of the channels permeable to both monovalent and divalent cations.