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The intrinsic mobility of intracellular H(+) ions was investigated by confocally imaging the longitudinal movement of acid inside rabbit ventricular myocytes loaded with the acetoxymethyl ester (AM) form of carboxy-seminaphthorhodafluor-1 (carboxy-SNARF-1). Acid was diffused into one end of the cell through a patch pipette filled with an isotonic KCl solution of pH 3.0. Intracellular H(+) mobility was low, acid taking 20-30 s to move 40 microm down the cell. Inhibiting sarcolemmal Na(+)-H(+) exchange with 1 mM amiloride had no effect on this time delay. Net H(+)(i) movement was associated with a longitudinal intracellular pH (pH(i)) gradient of up to 0.4 pH units. H(+)(i) movement could be modelled using the equations for diffusion, assuming an apparent diffusion coefficient for H(+) ions (D(H)(app)) of 3.78 x 10(-7) cm(2) s(-1), a value more than 300-fold lower than the H(+) diffusion coefficient in a dilute, unbuffered solution. Measurement of the intracellular concentration of SNARF (approximately 400 microM) and its intracellular diffusion coefficient (0.9 x 10(-7) cm(2) s(-1)) indicated that the fluorophore itself exerted an insignificant effect (between 0.6 and 3.3 %) on the longitudinal movement of H(+) equivalents inside the cell. The longitudinal movement of intracellular H(+) is discussed in terms of a diffusive shuttling of H(+) equivalents on high capacity mobile buffers which comprise about half (approximately 11 mM) of the total intrinsic buffering capacity within the myocyte (the other half being fixed buffer sites on low mobility, intracellular proteins). Intrinsic H(+)(i) mobility is consistent with an average diffusion coefficient for the intracellular mobile buffers (D(mob)) of ~9 x 10(-7) cm(2) s(-1).

Intracellular H(+) mobility was estimated in the rabbit isolated ventricular myocyte by diffusing HCl into the cell from a patch pipette, while imaging pH(i) confocally using intracellular ratiometric SNARF fluorescence. The delay for acid diffusion between two downstream regions approximately 40 microm apart was reduced from approximately 25 s to approximately 6 s by replacing Hepes buffer in the extracellular superfusate with a 5 % CO(2)/HCO(3)(-) buffer system (at constant pH(o) of 7.40). Thus CO(2)/HCO(3)(-) (carbonic) buffer facilitates apparent H(+)(i) mobility. The delay with carbonic buffer was increased again by adding acetazolamide (ATZ), a membrane permeant carbonic anhydrase (CA) inhibitor. Thus facilitation of apparent H(+)(i) mobility by CO(2)/HCO(3)(-) relies on the activity of intracellular CA. By using a mathematical model of diffusion, the apparent intracellular H(+) equivalent diffusion coefficient (D(H)(app)) in CO(2)/HCO(3)(-)-buffered conditions was estimated to be 21.9 x 10(-7) cm(2) s(-1), 5.8 times faster than in the absence of carbonic buffer. Facilitation of H(+)(i) mobility is discussed in terms of an intracellular carbonic buffer shuttle, catalysed by intracellular CA. Turnover of this shuttle is postulated to be faster than that of the intrinsic buffer shuttle. By regulating the carbonic shuttle, CA regulates effective H(+)(i) mobility which, in turn, regulates the spatiotemporal uniformity of pH(i). This is postulated to be a major function of CA in heart.

1. A fall of intracellular pH (pHi) typically depresses cardiac contractility. Among the many mechanisms underlying this depression, an inhibitory effect of acidosis upon the sarcoplasmic reticulum (SR) Ca2+ release channel has been predicted, but not so far demonstrated in the intact cardiac myocyte. In the present work, pHi was manipulated experimentally while confocal imaging was used to record spontaneous 'Ca2+ sparks' (local SR Ca2+ release events) in rat isolated myocytes loaded with the fluorescent Ca2+ indicator fluo-3. In other experiments, whole cell (global) pHi or [Ca2+]i was measured by microfluorimetry (using, respectively, intracellular carboxy SNARF-1 and indo-1). 2. Reducing pHi (i) increased whole cell intracellular [Ca2+] transients induced either electrically or by addition of caffeine, whereas (ii) it decreased spontaneous Ca2+ spark frequency. Conversely, raising pHi increased spontaneous Ca2+ spark frequency. 3. Blocking sarcolemmal Ca2+ influx with 10 mM Ni2+, or reducing external pH by 1.0 unit, had no effect on the pHi-dependent changes in spontaneous Ca2+ spark frequency. 4. Decreasing pHi over the range 7.78-7.20, decreased Ca2+ spark frequency exponentially as a function of pHi, with frequency declining by approximately 33 % for a 0.2 unit fall in pHi. In contrast, over the same pHi range, Ca2+ spark amplitude was unaffected. Intracellular acidosis produced a slight slowing of Ca2+ spark relaxation. 5. The results indicate that, in the intact myocyte, a reduced pHi decreases the probability of opening of the SR Ca2+ release channel. This phenomenon may contribute to the negative inotropic effects of acidosis.

This study describes the use of a microperfusion system to create rapid, large regional changes in intracellular pH (pH(i)) within single ventricular myocytes. The spatial distribution of pH(i) in single myocytes was measured with seminaphthorhodafluor-1 fluorescence using confocal imaging. Changes in pH(i) were induced by local external application of NH(4)Cl, CO(2), or sodium propionate. Local application was achieved by simultaneously directing two parallel square microstreams, each 275 microm wide, over a single myocyte oriented perpendicular to the direction of flow. One stream contained the control solution, and the other contained a weak acid or base. End-to-end, stable pH(i) gradients as large as 1 pH unit were readily created with this technique. This result indicates that pH within a single cardiac cell may not always be spatially uniform, particularly when weak acid or base gradients are present, which can occur, for example, in regional myocardial ischemia. The microperfusion method should be useful for studying the effects of localized acidosis on myocyte function, estimating intracellular ion diffusion rates, and, possibly, inducing regional changes in other important intracellular ions.

1. Intracellular pH was recorded fluorimetrically by using carboxy-SNARF-1, AM-loaded into superfused ventricular myocytes isolated from guinea-pig heart. Intracellular acid and base loads were induced experimentally and the changes of pHi used to estimate intracellular buffering power (beta). The rate of pHi recovery from acid or base loads was used, in conjunction with the measurements of beta, to estimate sarcolemmal transporter fluxes of acid equivalents. A combination of ion substitution and pharmacological inhibitors was used to dissect acid effluxes carried on Na+-H+ exchange (NHE) and Na+-HCO3- cotransport (NBC), and acid influxes carried on Cl--HCO3- exchange (AE) and Cl--OH- exchange (CHE). 2. The intracellular intrinsic buffering power (betai), estimated under CO2/HCO3--free conditions, varied inversely with pHi in a manner consistent with two principal intracellular buffers of differing concentration and pK. In CO2/HCO3--buffered conditions, intracellular buffering was roughly doubled. The size of the CO2-dependent component (betaCO2) was consistent with buffering in a cell fully open to CO2. Because the full value of betaCO2 develops slowly (2.5 min), it had to be measured under equilibrium conditions. The value of betaCO2 increased monotonically with pHi. 3. In 5 % CO2/HCO3--buffered conditions (pHo 7.40), acid extrusion on NHE and NBC increased as pHi was reduced, with the greater increase occurring through NHE at pHi < 6.90. Acid influx on AE and CHE increased as pHi was raised, with the greater increase occurring through AE at pHi > 7.15. At resting pHi (7.04-7.07), all four carriers were activated equally, albeit at a low rate (about 0.15 mM min-1). 4. The pHi dependence of flux through the transporters, in combination with the pHi and time dependence of intracellular buffering (betai + betaCO2), was used to predict mathematically the recovery of pHi following an intracellular acid or base load. Under several conditions the mathematical predictions compared well with experimental recordings, suggesting that the model of dual acid influx and acid efflux transporters is sufficient to account for pHi regulation in the cardiac cell. Key properties of the pHi control system are discussed.

1. The mechanism of pHi recovery from an intracellular alkali load (induced by acetate prepulse or by reduction/removal of ambient PCO2) was investigated using intracellular SNARF fluorescence in the guinea-pig ventricular myocyte. 2. In Hepes buffer (pHo 7.40), pHi recovery was inhibited by removal of extracellular Cl-, but not by removal of Na+o or elevation of K+o. Recovery was unaffected by the stilbene drug DIDS (4,4-diisothiocyanatostilbene-disulphonic acid), but was slowed dose dependently by the stilbene drug DBDS (dibenzamidostilbene-disulphonic acid). 3. In 5 % CO2/HCO3- buffer (pHo 7.40), pHi recovery was faster than in Hepes buffer. It consisted of an initial rapid recovery phase followed by a slow phase. Much of the rapid phase has been attributed to CO2-dependent buffering. The slow phase was inhibited completely by Cl-o removal but not by Na+o removal or K+o elevation. 4. At a test pHi of 7.30 in CO2/HCO3- buffer, the slow phase was inhibited 70 % by DIDS. The mean DIDS-inhibitable acid influx was equivalent in magnitude to the HCO3--stimulated acid influx. Similarly, the DIDS-insensitive influx was equivalent to that estimated in Hepes buffer. 5. We conclude that two independent sarcolemmal acid-loading carriers are stimulated by a rise of pHi and account for the slow phase of recovery from an alkali load. The results are consistent with activation of a DIDS-sensitive Cl--HCO3- anion exchanger (AE) to produce HCO3- efflux, and a DIDS-insensitive Cl--OH- exchanger (CHE) to produce OH- efflux. H+-Cl- co-influx as the alternative configuration for CHE is not, however, excluded. 6. The dual acid-loading system (AE plus CHE), previously shown to be activated by a fall of extracellular pH, is thus activated by a rise of intracellular pH. Activity of the dual-loading system is therefore controlled by pH on both sides of the cardiac sarcolemma.

1. Following an intracellular alkali load (imposed by acetate prepulsing in CO2/HCO3- buffer), intracellular pH (pHi) of the guinea-pig ventricular myocyte (recorded from intracellular SNARF fluorescence) recovers to control levels. Recovery has two phases. An initial rapid phase (lasting up to 2 min) is followed by a later slow phase (several minutes). Inhibition of sarcolemmal acid-loading carriers (by removal of extracellular Cl-) inhibits the later, slow phase but the initial rapid recovery phase persists. It also persists in the absence of extracellular Na+ and in the presence of the HCO3- transport inhibitor DIDS (4,4-di-isothiocyanatostilbene-2, 2-disulphonic acid). 2. The rapid recovery phase is not evident if the alkali load has been induced by reducing PCO2 (from 10 to 5 %), and it is inhibited in the absence of CO2/HCO3- buffer (i.e. Hepes buffer). It is also slowed by the carbonic anhydrase (CA) inhibitor acetazolamide (ATZ). We conclude that it is caused by buffering of the alkali load through the hydration of intracellular CO2 (CO2-dependent buffering). 3. The time course of rapid recovery is consistent with an intracellular CO2 hydration rate constant (k1) of 0.36 s-1 in the presence of CA activity, and 0.14 s-1 in the absence of CA activity. This latter k1 value matches the literature value for uncatalysed CO2 hydration in free solution. Natural CO2 hydration is accelerated 2.6-fold in the ventricular myocyte by endogenous CA. 4. The rapid recovery phase represents a period when the intracellular CO2/HCO3- buffer is out of equilibrium (OOE). Modelling of the recovery phase using our k1 value, indicates that OOE conditions will normally extend for at least 2 min following a step rise in pHi (at constant PCO2). If CA is inactive, this period can be as long as 5 min. During normal pHi regulation, the recovery rate during these periods cannot be used as a measure of sarcolemmal acid loading since it is a mixture of slow CO2-dependent buffering and transmembrane acid loading. The implication of this finding for quantification of pHi regulation during alkalosis is discussed.

1. Intracellular pH (pHi) was recorded in single, isolated guinea-pig ventricular myocytes using the pH-sensitive fluorophore, carboxy-SNARF-1 (AM-loaded). 2. The dual acid extrusion system in this cell (Na(+)-H+ antiport and Na(+)-HCO3- symport) was activated by inducing an intracellular acid load, produced by addition and subsequent removal of extracellular 10 mM NH4Cl. Under these conditions, it is known that both acid-equivalent extruders are activated about equally. 3. Application of phenylephrine (100 microM; alpha-adrenergic agonist) resulted in an inhibition of pHi recovery from an acid load, recorded in HCO3-buffered medium containing 1.5 mM amiloride (amiloride inhibits Na(+)-H+ antiport; under these conditions pHi recovery is mediated through only the Na(+)-HCO3- symport carrier). This inhibitory effect of phenylephrine was prevented by the alpha 1-antagonist, prazosin (0.1 microM) and was unaffected by propranolol (1 microM). 4. Application of phenylephrine in Hepes-buffered medium (only Na(+)-H+ antiport is active under these conditions) elicited a stimulation of pHi recovery, again prevented by prazosin (0.1 microM). 5. These results point to an alpha 1 inhibition of Na(+)-HCO3- symport and an alpha 1 stimulation of Na+-H+ antiport. 6. Both adrenaline (1-5 microM) and noradrenaline (5 microM) slowed pHi recovery recorded in HCO3(-)-buffered solution containing amiloride (1.5 mM). The similarity of this result with that obtained previously using phenylephrine (paragraph 3) suggests that all three agonists inhibit the Na(+)-HCO3- symport through alpha 1 activation. 7. Isoprenaline (1 microM; beta-adrenergic agonist) slowed pHi recovery in Hepes-buffered solution but stimulated recovery in a HCO3(-)-buffered solution containing amiloride (1.5 mM). These results suggest that beta activation slows Na(+)-H+ antiport but stimulates Na(+)-HCO3- symport. 8. When both acid-equivalent extrusion carriers were inhibited in Na(+)-free, HCO3(-)-buffered medium, phenylephrine or isoprenaline had no effect on pHi, ruling out any effect of the adrenergic agonists on background acid-loading mechanisms. 9. Under physiological conditions (CO2/HCO3(-)-buffered solution, no amiloride), when both acid extruders would be activated by an intracellular acid load, application of phenylephrine, adrenaline or noradrenaline were found to slow pHi recovery. In contrast, isoprenaline stimulated pHi recovery under the same conditions.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Intracellular pH (pHi) was recorded in isolated guinea-pig ventricular myocytes using the pH-sensitive fluoroprobe, carboxy-SNARF-1 (carboxy-seminaphthorhodafluor). 2. Addition and removal of 10 mM NH4Cl was used to induce an intracellular acid load in a myocyte perfused with HCO3(-)-buffered solution containing amiloride. Under these conditions, subsequent pHi recovery is known to rely upon Na(+)-HCO3- co-transport into the cell. The application of 0.5-5 microM adrenaline resulted in an inhibition of this pHi recovery. 3. In HEPES-buffered solution, where acid extrusion is mediated primarily by Na(+)-H+ antiport, pHi recovery from an acid load was stimulated by the application of adrenaline. 4. In HCO3-/CO2-buffered solution (no amiloride), when both acid-aquivalent extruders are activated by an intracellular acidification, adrenaline was found to slow pHi recovery. 5. When both carriers were inhibited in Na(+)-free, HCO3(-)-buffered medium, adrenaline had no effect on pHi, ruling out any effect of the catecholamine on background acid loading. 6. The voltage clamp technique was used to test if the inhibitory effect of adrenaline on amiloride-resistant, HCO3(-)-dependent pHi recovery was due to an efflux of HCO3- ions through catecholamine-activated anion channels. During pHi recovery, membrane depolarization, sufficient to reverse the electrochemical driving force acting on HCO3-, had no effect upon pHi recovery rate. 7. The above results show that adrenaline has direct but opposite effects on Na(+)-HCO3- co-transport and Na(+)-H+ antiport. In the presence of this agonist, the pHi dependence of Na(+)-HCO3- symport was shifted to the left along the pHi axis by 0.13 +/- 0.03 units (n = 4) whereas that for Na(+)-H+ antiport was shifted in the opposite direction by only 0.07 +/- 0.01 units (n = 3). Following an acid load, the net effect of adrenaline under physiological conditions was, therefore, a slowing of pHi recovery. 8. The application of extracellular ATP (ATPo, 10-50 microM) mimicked the effects of adrenaline on both Na(+)-H+ exchange and Na(+)-HCO3- symport. 9. Adenosine (50 microM) and ADP (50 microM) did not induce any inhibition of Na(+)-HCO3- symport, suggesting that the inhibition induced by ATP was not mediated through P1 or P2-purinergic receptors. 10. We conclude that Na(+)-H+ antiport and Na(+)-HCO3- symport are both coupled to adrenaline and ATPo receptors. Activation of these receptors switches acid-equivalent extrusion from a situation dependent on both HCO3- and H+ ions to one nearly exclusively dependent upon H+.

1. Intracellular pH (pHi) was recorded ratiometrically in isolated guinea-pig ventricular myocytes using the pH-sensitive fluoroprobe, carboxy-SNARF-1 (carboxy-seminaphthorhodafluor). 2. Following an intracellular acid load (10 mM NH4 Cl removal), pHi recovery in HEPES-buffered Tyrode solution was inhibited by 1.5 mM amiloride (Na(+)-H+ antiport blocker). In the presence of amiloride, switching from HEPES buffer to HCO3-/CO2 (pHo of both solutions = 7.4) stimulated a pHi recovery towards more alkaline levels. 3. Amiloride-resistant, HCO(3-)-dependent pHi recovery was inhibited by removal of external Na+ (replaced by N-methyl-D-glucamine), whereas removal of external Cl- (replaced by glucuronate, leading to depletion of internal Cl-), removal of external K+, or decreasing external Ca2+ by approximately tenfold had no inhibitory effect. These results suggest that the amiloride-resistant recovery is due to a Na(+)-HCO3- cotransport into the cell. 4. The stilbene derivative DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid, 500 microM) slowed Na(+)-HCO(3-)-dependent pHi recovery. 5. Intracellular pH increased in Cl(-)-free solution and this increase still occurred in Na(+)-free solution indicating that it is not caused via Na(+)-HCO3- symport and is more likely to be due to Cl- efflux in exchange for HCO3- influx on a sarcolemmal Cl(-)-HCO3- exchanger. The lack of any significant pHi recovery from intracellular acidosis in Na(+)-free solution suggests that this exchanger does not contribute to acid-equivalent extrusion. 6. Possible voltage sensitivity and electrogenicity of the co-transport were examined by using the whole-cell patch clamp technique in combination with SNARF-1 recordings of pHi. Stepping the holding potential from -110 to -40 mV did not affect amiloride-resistant pHi recovery from acidosis. Moreover, following an intracellular acid load, the activation of Na(+)-HCO3- co-influx (by switching from HEPES to HCO3-/CO2 buffer) produced no detectable outward current (outward current would be expected if the coupling of HCO3- with Na+ were > 1.0). 7. Intracellular intrinsic buffering power (beta i) was assessed as a function of pHi (beta i computed from the decrease of pHi following reduction of extracellular NH4 Cl in amiloride-containing solution). beta i in the ventricular myocyte increases roughly linearly with a decrease in pHi according the following equation: beta i = -28(pHi) +222.6. 8. Comparison of acid-equivalent efflux via Na(+)-HCO3- symport and Na(+)-H+ antiport showed that, following an intracellular acidosis, the symport accounts for about 40% of total acid efflux, the other 60% being carried by the antiport.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Intracellular pH was recorded (double-barrelled pH-selective microelectrodes) in single ventricular myocytes and whole papillary muscles isolated from guinea-pig heart. Both preparations were acid-loaded by various manoeuvres (addition and removal of external NH4Cl or CO2) in order that a comparison could be made of the size and speed of intracellular pH changes and hence of the apparent intracellular buffering power (beta). 2. For the same acid-loading procedure, the size of intracellular pH (pHi) changes was about threefold larger in the isolated myocyte than in whole papillary muscle. The rate of initial acid loading as well as the subsequent rate of pHi recovery (caused by acid extrusion from the cell) were also threefold faster in the myocyte. Estimates of apparent intrinsic (non-CO2) buffering power, based upon the size of pHi changes during acid loading, were 15-20 mmol l-1 for the myocyte and about 70 mmol l-1 for whole muscle. This latter value is similar to previous estimates of beta in heart. 3. When acid extrusion was reduced by applying a high dose of amiloride (1 mmol l-1), then the size of the pHi change during acid loading increased greatly in papillary muscle but changed much less in the myocyte; beta now appeared to be about 30 mmol l-1 in whole muscle but remained essentially unchanged in the myocyte. 4. We conclude that previous values for beta in cardiac muscle have been greatly overestimated because of the presence of sarcolemmal acid extrusion. Paradoxically, this error in estimating beta is far less evident in the isolated myocyte. We suggest that this is because a much more rapid acid loading is achievable in the myocyte so that acid loading will be blunted less by acid extrusion than in whole muscle. We present a simple mathematical model that demonstrates this phenomenon. We conclude that beta in ventricular muscle is likely to resemble that measured in the isolated myocyte, i.e. 15-20 mmol l-1. 5. Slow acid loading in whole ventricular muscle will also affect the kinetics of pHi changes. The model indicates that the rate of pHi recovery from an acid load in papillary muscle does not reflect the pHi dependence of acid extrusion. Instead, it is heavily influenced by the slow rate of acid loading. This emphasises that great care should be taken when interpreting the kinetics of pHi changes in multicellular ventricular preparations.

Intracellular pH (pHi) in sheep cardiac Purkinje fibres is controlled by sarcolemmal Na+/H+ and Cl-/HCO3- exchange. At normal pHo (7.4), Na+/H+ exchange mediates an acid efflux whenever pHi falls and Cl-/HCO3- exchange mediates an equivalent acid influx in response to a rise in pHi. Intracellular pH is also influenced by Ca2+i, which can activate force development leading to the anaerobic production of lactic acid. This is evident after an increase in stimulation rate which reversibly reduces both pHi and extracellular surface pH (pHs). Rate-dependent pHi changes are inhibited following inhibition of glycolysis, indicating that they are caused by accumulation of lactic acid. In some cases, the efflux of lactic acid may provide a faster method for recovery of pHi from a metabolic acidosis than that provided by Na+/H+ exchange. Finally, direct pHi measurement in isolated mammalian ventricular myocytes suggests that the intrinsic intracellular buffering power (beta) of ventricular tissue may be considerably lower than previously believed. An accurate knowledge of beta is essential for calculating net membrane fluxes of acid equivalents from changes in pHi.

Intracellular pH (pHi) and Na (aina) were recorded in isolated sheep cardiac Purkinje fibres using ion-selective microelectrodes while simultaneously recording twitch tension. A fall of pHi stimulated acid-extrusion via sarcolemmal Na-H exchange but the extrusion was inhibited by reducing extracellular pH (pHo), indicating an inhibitory effect of external H ions upon the exchanger. Intracellular acidosis can reduce contraction by directly reducing myofibrillar Ca2+ sensitivity. The activation of Na-H exchange at low pHi can offset this direct inhibitory effect of H+ ions since exchange-activation elevates aina which then indirectly elevates Ca2+i (via Na-Ca exchange) thus tending to restore tension. This protection of contraction during intracellular acidosis can be removed if extracellular pH is also allowed to fall since, under these conditions, Na-H exchange is inhibited.

Intracellular Na and pH were measured with recessed-tip ion-selective microelectrodes in voltage-clamped sheep cardiac Purkinje fibers. Intracellular Na activity (aiNa) was elevated by inhibiting the Na/K pump. This produced an increase of twitch tension that had a steep dependence on the increase of aiNa. These effects of aiNa on twitch tension are probably mediated by an Na-Ca exchange. An increase of aiNa also produced a component of tonic tension that appears to be produced directly by the Na-Ca exchange. The dependence of tonic tension and aiNa on membrane potential suggests that this exchange process may be voltage-sensitive. The increase of aiNa is associated with an intracellular acidification that appears to be secondary to an increase of [Ca2+]i produced by Na-Ca exchange. Therefore, as well as affecting [Ca2+]i, Na-Ca exchange can under some circumstances influence pHi indirectly, and this complicates the interpretation of changes in tension, since protons and Ca ions have opposite effects on contractile force.

Changes of intracellular Ca concentration in cardiac muscle have significant effects on transmembrane currents and many of these effects can be accounted for by postulating the existence of Ca-activated K channels in the heart. However, the evidence that such channels exist is equivocal. This is partly because of technical problems, for example the difficulty of identifying an individual ionic current amongst the many currents that exist in the heart. An additional problem, however, is posed by the fact that other currents may also be modulated by Ca ions. It is important therefore to distinguish between these currents and those caused by Ca-activated, K-specific channels. In this review we consider the evidence for Ca activated currents in the heart and, in particular, we discuss whether or not these currents are carried exclusively by K ions.