The relationship among intracellular sodium activity, calcium, and strophanthidin inotropy in canine cardiac Purkinje fibers

The role of sodium and calcium ions in strophanthidin inotropy was studied by measuring simultaneously the electrical, mechanical, and intracellular sodium ion activities in electrically driven cardiac Purkinje fibers under conditions that change the intracellular sodium or calcium level (tetrodotoxin, strophanthidin, high calcium, and norepinephrine). Tetrodotoxin (TTX; 1-5 X 10(-6)M) shifted the action potential plateau to more negative values, shortened the action potential duration, and decreased the contractile tension and the intracellular sodium ion activity (aiNa). The changes in tension and in aiNa caused by TTX appear to be related since they had similar time courses. Strophanthidin (2-5 X 10(-7)M) increased tension and aiNa less in the presence of TTX, and, for any given value of aiNa, tension was less than in the absence of TTX. Increasing extracellular calcium (from 1.8 to 3.3-3.6 mM) or adding norepinephrine (0.5-1 X 10(-6)M) increased tension and decreased aiNa less in the presence than in the absence of TTX. When two of the above procedures were combined, the results were different. Thus, during the increase in aiNa and tension caused by strophanthidin in the presence of TTX, increasing calcium or adding norepinephrine increased tension markedly but did not increase aiNa further. In a TTX-high calcium or TTX-norepinephrine solution, adding strophanthidin increased both tension and aiNa, and the increase in tension was far greater than in the presence of TTX alone. The results indicate that: (a) the contractile force in Purkinje fibers is affected by a change in aiNa; (b) a decrease in aiNa by TTX markedly reduces the inotropic effect of strophanthidin, possibly as a consequence of depletion of intracellular calcium; (c) increasing calcium influx with norepinephrine or high calcium in the TTX-strophanthidin solution produces a potentiation of tension development, even if aiNa does not increase further; and (d) when the calcium influx is already increased by high calcium or norepinephrine, strophanthidin has its usual inotropic effect even in the presence of TTX. In conclusion, the positive inotropic effect of strophanthidin requires that an increase in aiNa be associated with suitable calcium availability.     

Ionic mechanisms of pacemaker activity in cardiac Purkinje fibers.

Rhythmic activity in cardiac Purkinje fibers can be analyzed by using the voltage clamp technique to study pacemaker currents. In normally polarized preparations, pacemaker activity can be generated by two distinct ionic mechanisms. The standard pacemaker potential (phase 4 depolarization) involves a slow potassium current, IK2. Following action potential repolarization, the IK2 channels slowly deactivate and thus unmask a steady background inward current. The resulting net inward current causes the slow pacemaker depolarization. Epinephrine accelerates the diastolic depolarization by promoting more complete and more rapid deactivation of IK2 over the pacemaker range of potentials. The catecholamine acts rather selectively on the voltage dependence of the gating mechanism, without altering the basic character of the pacemaker process. The nature of the pacemaker depolarization is altered by intoxication with high concentrations of cardiac glycosides or aglycones. These compounds promote spontaneous impulses in Purkinje fibers by a mechanism that supersedes the ordinary IK2 pacemaker. The digitalis-induced depolarization is generated by a transient inward current that is either absent or very small in untreated preparations. The transient inward current is largely carried by sodium ions. Its unusual time course probably reflects an underlying subcellular event, the oscillatory release of calcium ions from intracellular stores.     

Electrogenic sodium extrusion in cardiac Purkinje fibers

Thin canine cardiac Purkinje fibers in a fast flow chamber were exposed to K-free fluid for 15 s to 6 min to initiate "sodium loading," then returned to K-containing fluid to stimulate the sodium pump. The electrophysiological effects of enhanced pump activity may result from extracellular K depletion caused by enhanced cellular uptake of K or from an increase in the current generated as a result of unequal pumped movements of Na and K, or from both. The effects of pump stimulation were therefore studied under three conditions in which lowering the external K concentration ([K]0) causes changes opposite to those expected from an increase in pump current. First, the resting potential of Purkinje fibers may have either a "high" value of a "low" (less negative) value: at the low level of potential, experimental reduction of [K]0 causes depolarization, whereas an increase in pump current should cause hyperpolarization. Second, in regularly stimulated Purkinje fibers, lowering [K]0 prolongs the action potential, whereas an increase in outward pump current should shorten it. Finally, lowering [K]0 enhances spontaneous "pacemaker" activity in Purkinje fibers, whereas an increase in outward pump current should reduce or abolish spontaneous activity. Under all three conditions, we find that the effects of temporary stimulation of the sodium pump are those expected from a transient increase in outward pump current, not those expected from K depletion.     

Intracellular sodium ion activity: reliable measurement and stimulation-induced change in cardiac Purkinje fibers

Recently Na+-selective microelectrodes (NaSM) have been used to measure quantitatively small changes in intracellular sodium ion activity (aiNa) and to determine a precise time course of comparatively rapid change in aiNa. In such studies, accurate measurement of aiNa requires the following criteria: (i) NaSM should have a fast response time and (ii) an NaSM and a conventional voltage microelectrode should measure the same membrane potential. These criteria were evaluated by measuring aiNa when membrane potential of cardiac Purkinje fibers was suddenly hyperpolarized and depolarized by changing stimulation rate. The NaSM coated with a conductive silver paint had fast response times so that rapid changes in aiNa could be reliably measured. The cardiac Purkinje fibers stimulated at a constant rate generated uniform membrane voltage and the NaSM and conventional microelectrode measured virtually the same membrane potential. This result is somewhat different from that reported under voltage-clamp condition by other investigators. The aiNa of the fibers increased as the stimulation rate was increased over the range of 0.5-3 Hz. In fibers stimulated at 1 Hz, cessation of stimulation was immediately followed by an exponential decline of aiNa with an average time constant of 53 +/- 9 s (SD, n = 8), or rate constant of 0.020 +/- 0.004/s. Restimulation of the fibers produced an exponential rise of aiNa with an average time constant of 65 +/- 12 s (n = 8). Similar results were obtained in fibers stimulated at 2 Hz.(ABSTRACT TRUNCATED AT 250 WORDS)     

The pacemaker current in cardiac Purkinje myocytes

It is generally assumed that in cardiac Purkinje fibers the hyperpolarization activated inward current i(f) underlies the pacemaker potential. Because some findings are at odds with this interpretation, we used the whole cell patch clamp method to study the currents in the voltage range of diastolic depolarization in single canine Purkinje myocytes, a preparation where many confounding limitations can be avoided. In Tyrode solution ([K+]o = 5.4 mM), hyperpolarizing steps from Vh = -50 mV resulted in a time-dependent inwardly increasing current in the voltage range of diastolic depolarization. This time- dependent current (iKdd) appeared around -60 mV and reversed near EK. Small superimposed hyperpolarizing steps (5 mV) applied during the voltage clamp step showed that the slope conductance decreases during the development of this time-dependent current. Decreasing [K+]o from 5.4 to 2.7 mM shifted the reversal potential to a more negative value, near the corresponding EK. Increasing [K+]o to 10.8 mM almost abolished iKdd. Cs+ (2 mM) markedly reduced or blocked the time-dependent current at potentials positive and negative to EK. Ba2+ (4 mM) abolished the time-dependent current in its usual range of potentials and unmasked another time-dependent current (presumably i(f)) with a threshold of approximately -90 mV (> 20 mV negative to that of the time-dependent current in Tyrode solution). During more negative steps, i(f) increased in size and did not reverse. During i(f) the slope conductance measured with small (8-10 mV) superimposed clamp steps increased. High [K+]o (10.8 mM) markedly increased and Cs+ (2 mM) blocked i(f). We conclude that: (a) in the absence of Ba2+, a time-dependent current does reverse near EK and its reversal is unrelated to K+ depletion; (b) the slope conductance of that time-dependent current decreases in the absence of K+ depletion at potentials positive to EK where inactivation of iK1 is unlikely to occur. (c) Ba2+ blocks this time-dependent current and unmasks another time-dependent current (i(f)) with a more negative (> 20 mV) threshold and no reversal at more negative values; (d) Cs+ blocks both time-dependent currents recorded in the absence and presence of Ba2+. The data suggest that in the diastolic range of potentials in Purkinje myocytes there is a voltage- and time-dependent K+ current (iKdd) that can be separated from the hyperpolarization- activated inward current i(f).     

Role of intracellular sodium activity in the control of contraction in cardiac purkinje fibers

The role of intracellular sodium activity (a_Naⁱ) in the control of force was studied in sheep cardiac Purkinje fibers exposed to norepinephrine (NE) and high [Ca]_o in the absence and presence of overdrive or of a low concentration of strophanthidin. Both NE and high [Ca]_o decrease a_Naⁱ and increase force, while overdrive increases and low strophanthidin decreases both parameters. In the presence of NE, overdrive increases a_Naⁱ less than force and is followed by a more pronounced undershoot in a_Naⁱ and force. In contrast, in high [Ca]_o overdrive increases a_Naⁱ more than force and is followed by a less pronounced undershoot in a_Naⁱ and force than in NE. High [Ca]_o increases force to a peak, but then the decreasing a_Naⁱ reduces force. In all these conditions, a_Naⁱ determines force changes during recovery from overdrive. NE and high [Ca]_o decrease a_Naⁱ less and increase force more in low strophanthidin. Thus, changes in a_Naⁱ modulate the increase in force due to increased Ca influx and control force development when Ca influx is either unchanged (low strophanthidin) or has reached a steady state (high [Ca]_o, recovery from overdrive).     

Fluctuations in membrane current driven by intracellular calcium in cardiac Purkinje fibers

Spontaneous oscillatory fluctuations in membrane potential are often observed in heart cells, but their basis remains controversial. Such activity is enhanced in cardiac Purkinje fibers by exposure to digitalis or K-free solutions. Under these conditions, we find that voltage noise is generated by current fluctuations that persist when membrane potential is voltage clamped. Power spectra of current signals are not made up of single time-constant components, as expected from gating of independent channels, but are dominated by resonant characteristics between 0.5 and 2 HZ. Our evidence suggests that the periodicity arises from oscillatory variations in intracellular free Ca that control ion movements across the surface membrane. The current fluctuations are strongly cross-correlated with oscillatory fluctuations in contractile force, and are inhibited by removing extracellular Ca or exposure to D600. Chelating intracellular Ca with injected EGTA also abolishes the current fluctuations. The oscillatory mechanism may involve cycles of Ca (or Sr) movement between sarcoplasmic reticulum and myoplasm, as previously suggested for skinned cardiac preparations. Our experiments in intact cells indicate that changes in surface membrane potential can modulate cytoplasmic Ca oscillations in frequency and perhaps amplitude as well. A two-way interaction between surface membrane potential and intracellular Ca stores may be a common feature of heart, neuron, and other cell types.     

Relation between intracellular Na ion activity and tension of sheep cardiac Purkinje fibers exposed to dihydro-ouabain.

The intracellular Na ion activity (aiNa) and the contractile tension (T) of sheep cardiac Purkinje fibers were simultaneously measured employing recessed-tip Na+-selective glass microelectrodes and a mechano-electric transducer. The aiNa of 6.4 +/- 1.6 mM (mean +/- SD, n = 56) was obtained in fibers perfused with normal Tyrode's solution. The changes in aiNa and T were measured during and after the exposure of fibers to a cardiac glycoside, dihydro-ouabain (DHO) in concentrations between 5 X 10(-8) M and 10(-5) M. The exposure time to DHO was 15 min. Both aiNa and T did not change in fibers exposed to 5 X 10(-8) M DHO, and the threshold concentration for the effect of DHO appeared to be around 10(-7) M. In DHO concentrations greater than the threshold, the increases in aiNa and T strongly correlated during the onset of DHO effects. The recoveries of aiNa and T were variable and slow, being dependent on the DHO concentration. In those fibers which recovered from the effects of DHO, the time-course of aiNa recovery was similar to that of T recovery. In fibers exposed to DHO of 5 X 10(-6) M or greater, the apparent toxic effects were observed in both action potential and contraction after an initial increase in T. The fibers manifesting the apparent toxic effects has a aiNa of approximately 30 mM or greater. The results of this study indicate that the increase in aiNa is associated with the positive inotropic action of the cardiac glycoside.     

Effects of changes of intracellular pH on contraction in sheep cardiac Purkinje fibers

Intracellular pH (pHi) was measured with a pH-sensitive microelectrode in voltage-clamped sheep cardiac Purkinje fibers while tension was simultaneously measured. All solutions were nominally CO2/HCO3 free and were buffered with Tris. The addition of NH4Cl (5-20 mM) produced an initial intracellular alkalosis that was associated with an increase of twitch tension. At the same time, a component of voltage-dependent tonic tension developed. Prolonged exposure (greater than 5 min) to NH4Cl resulted in a slow recovery of pHi accompanied by a decrease of tension. Removal of NH4Cl produced a transient acidosis that was accompanied by a fall of force. In some experiments, there was then a transient recovery of force. If extracellular pH (pHo) was decreased, then pHi decreased slowly. Tension also fell slowly. An increase of pHo produced a corresponding increase of both force and pHi. The application of strophanthidin (10 microM) increased force and produced an intracellular acidosis. The addition of NH4Cl, to remove this acidosis partially, produced a significant increase of force. The above results show that contraction is sensitive to changes of intracellular but not extracellular pH. This pH dependence will therefore modify the contractile response to inotropic maneuvers that also affect pHi.     

Cesium,Na+-K+ pump and pacemaker potential in cardiac purkinje fibers

The mechanisms of the hyperpolarizing and depolarizing actions of cesium were studied in cardiac Purkinje fibers perfused in vitro by means of a microelectrode technique under conditions that modify either the Na⁺-K⁺ pump activity or I_f. Cs⁺ (2 mM) inconsistently increased and then decreased the maximum diastolic potential (MDP); and markedly decreased diastolic depolarization (DD). Increase and decrease in MDP persisted in fibers driven at fast rate (no diastolic interval and no activation of I_f). In quiescent fibers, Cs⁺ caused a transient hyperpolarization during which elicited action potentials were followed by a markedly decreased undershoot and a much reduced DD. In fibers depolarized at the plateau in zero [K⁺]_o (no I_f), Cs⁺ induced a persistent hyperpolarization. In 2 mM [K⁺]_o, Cs⁺ reduced the undershoot and suppressed spontaneous activity by hyperpolarizing and thus preventing the attainment of the threshold. In 7 mM [K⁺]_o, DD and undershoot were smaller and Cs⁺ reduced them. In 7 and 10 mM [K⁺]_o, Cs⁺ caused a small inconsistent hyperpolarization and a net depolarization in quiescent fibers; and decreased MDP in driven fibers. In the presence of strophanthidin, Cs⁺ hyperpolarized less. Increasing [Cs⁺]_o to 4, 8 and 16 mM gradually hyperpolarized less, depolarized more and abolished the undershoot. We conclude that in Purkinje fibers Cs⁺ hyperpolarizes the membrane by stimulating the activity of the electrogenic Na⁺-K⁺ pump (and not by suppressing I_f); and blocks the pacemaker potential by blocking the undershoot, consistent with a Cs⁺ block of a potassium pacemaker current.     

Membrane currents following activity in canine cardiac Purkinje fibers.

Recent experiments in canine Pukinje fibers (Gadsby and Cranefield, 1979) have shown that following a period of sodium loading in K+-free solution a slowly decaying outward current is observed. This current has been attributed to the activity of the electrogenic Na+-K+ exchange pump. In the present paper we show that similar slowly decaying outward currents are observed following prolonged periods of overdrive with action potentials or with brief depolarizing voltage clamp pulses. The dependent of the prolonged outward current on the duration and frequency of the preceding period of overdrive and on the potential following overdrive is reported. We also present results which indicate that a large portion of this current can be induced by phasic Na+ loading through the fast-inward channel.     

The effects of acidosis and bicarbonate on action potential repolarization in canine cardiac Purkinje fibers

Studies were performed on canine cardiac Purkinje fibers to evaluate the effects of acidosis and bicarbonate (HCO3) on action potential repolarization. Extracellular pH (pHe) was reduced from 7.4 to 6.8 by increasing carbon dioxide (CO2) concentration from 4 to 15% in a HCO3-buffered solution or by NaOH titration in a Hepes-buffered solution. Both types of acidosis produced a slowing of the rate of terminal repolarization (i.e., period of repolarization starting at about -60 mV and ending at the maximum diastolic potential) with an attendant increase in action potential duration of 10--20 ms. This was accompanied by a reduction in the maximum diastolic potential of 2--8 mV. In contrast, if the same pH change was made by keeping CO2 concentration constant and lowering extracellular HCO3 from 23.7 to 6.0 mM, in addition to the slowing of terminal repolarization, the plateau was markedly prolonged resulting in an additional 50- to 80-ms increase in action potential duration. If pHe was held constant at 7.4 and HCO3 reduced from 23.7 mM to 0 (Hepes-buffered solution), the changes in repolarization were nearly identical to those seen in 6.0 mM HCO3 except that terminal repolarization was unchanged. This response was unaltered by doubling the concentration of Hepes. Reducing HCO3 to 12.0 mM produced changes in repolarization of about one-half the magnitude of those in 6.0 mM HCO3. These findings suggest that in Purkinje fibers, HCO3 either acts as a current that slows repolarization or modulates the ionic currents responsible for repolarization.     

Effect of strophanthidin on intracellular Na ion activity and twitch tension of constantly driven canine cardiac Purkinje fibers.

Intracellular Na ion activity (aiNa) and twitch tension (T) of constantly driven (1 Hz) canine cardiac Purkinje fibers were measured simultaneously and continuously with neutral carrier Na+-selective microelectrodes and a force transducer. The aiNa of 8.9 +/- 1.4 mM (mean +/- SD, n = 52) was obtained in the driven fibers perfused with normal Tyrode solution. Temporary interruption of stimulation showed that aiNa of the driven fibers was approximately 1.5 mM greater than that of quiescent fibers. The constantly driven fibers were exposed to strophanthidin of 10(-8), 5 X 10(-8), 10(-7), 5 X 10(-7), and 10(-6) M for 5 min. No detectable changes in aiNa and T were observed in the fibers exposed to 10(-8) M strophanthidin, and the threshold concentration of the strophanthidin effect appeared to be approximately 5 X 10(-8) M. With concentrations greater than 5 X 10(-8) M, strophanthidin produced dose-dependent increases in aiNa and T. An increase in aiNa always accompanied an increase in T and after strophanthidin exposure both aiNa and T recovered completely. During onset and recovery periods of the strophanthidin effect the time course of change in aiNa was similar to that of change in T. A plot of T vs. aiNa during the onset and recovery periods showed a linear relationship between T and aiNa. These results indicate strongly that the positive inotropic effect of strophanthidin is closely associated with the increase in aiNa. Raising [K+]0 from 5.4 to 10.8 mM produced decreases in aiNa and T, and restoration of [K+]0 resulted in recoveries of aiNa and T. During the changes of [K+]0 the time course of change in aiNa was similar to that of the change in T. A steady-state sarcoplasmic Ca ion activity (aiCa) of 112 +/- 31 nM (mean +/- SD, n = 17) was obtained in the driven fibers with the use of neutral carrier Ca2+-selective microelectrodes. Temporary interruption produced 10-30% decreases in aiCa. No detectable changes in aiCa were observed in the fibers exposed to strophanthidin of 10(-7) M or less; 5 X 10(-7) and 10(-6) M strophanthidin produced 1.3-1.6 and 2-3-fold increases in aiCa, respectively. This result is consistent with the hypothesis that an increase in aiNa produces an increase in aiCa, which enhances Ca accumulation in the intracellular stores.     

Voltage-dependent block by tetrodotoxin of the sodium channel in rabbit cardiac Purkinje fibers.

The two-microelectrode, voltage-clamp technique was applied to rabbit cardiac Purkinje fibers to study the interaction of tetrodotoxin (TTX) with the slowly inactivating Na current. Binding of TTX to rested, inactivated, and activated channels was estimated by measuring the relative decrease of current at the beginning (rested and inactivated channels) and the end (activated channels) of a 1 s depolarizing clamp to -45 mV. The accelerated decline of the Na current in the presence of a submaximal dose of TTX was interpreted as an increase in blocking efficiency upon depolarization. The experiments show that activated as well as inactivated channels are more sensitive to TTX than are rested channels. The dissociation equilibrium constants for the three states are 3.5 X 10(-6) M for the rested, 0.94 X 10(-6) M for the activated, and 0.75 X 10(-6) M for the inactivated channels. The time course of activation block was dependent on TTX concentration. Rate constants for association and dissociation of the activated state are 1.3 X 10(6) M-1 X s-1 and 1.5 s-1, respectively.     

Strophanthidin inotropy in cardiac Purkinje fibers under conditions that vary cellular sodium or calcium

The role of sodium and calcium on strophanthidin inotropy was studied in canine cardiac Purkinje fibers perfused in vitro under conditions that vary cellular sodium and calcium. With high concentrations of strophanthidin (greater than or equal to 10(-7) M), force increases more in the presence of low [Ca]0 or high [Na]0 and less in the presence of a low sodium-calcium concentration solution than in Tyrode solution. In a solution with a low concentration of sodium-calcium containing strophanthidin, restoring [Na]0 to normal decreases and then re-increases force: when [Na]0 is decreased again, the force transiently overshoots. These effects of strophanthidin are exaggerated by metabolic inhibitors. In a low [Ca] solution, low concentrations of strophanthidin (3 X 10(-8) or 5 X 10(-8) M) re-increase force a little or not at all. On recovery, the transient force increase is not exaggerated by low strophanthidin and is absent after manganese exposure. The inotropy of low concentrations of strophanthidin is potentiated by norepinephrine, high [Ca]0 (4 mM), or by lowering [Na]0. Thus, the present results suggest that the inotropic action of high strophanthidin concentrations depends primarily on sodium and secondarily on calcium, and that the inotropic action of low concentrations of strophanthidin involves a modification of the cell response to calcium.     

Calcium- and voltage-activated plateau currents of cardiac Purkinje fibers

We have used the two-microelectrode voltage-clamp technique to investigate the components of membrane current that contribute to the formation of the early part of the plateau phase of the action potential of calf cardiac Purkinje fibers. 3,4-Diaminopyridine (50 microM) reduced the net transient outward current elicited by depolarizations to potentials positive to -30 mV but had no consistent effect on contraction. We attribute this effect to the blockade of a voltage-activated transient potassium current component. Ryanodine (1 microM), an inhibitor of sarcoplasmic reticulum calcium release and intracellular calcium oscillations in Purkinje fibers (Sutko, J.L., and J.L. Kenyon. 1983. Journal of General Physiology. 82:385-404), had complex effects on membrane currents as it abolished phasic contractions. At early times during a depolarization (5-30 ms), ryanodine reduced the net outward current. We attribute this effect to the loss of a component of calcium-activated potassium current caused by the inhibition of sarcoplasmic reticulum calcium release and the intracellular calcium transient. At later times during a depolarization (50-200 ms), ryanodine increased the net outward current. This effect was not seen in low-sodium solutions and we could not observe a reversal potential over a voltage range of -100 to +75 mV. These data suggest that the effect of ryanodine on the late membrane current is attributable to the loss of sodium-calcium exchange current caused by the inhibition of sarcoplasmic reticulum calcium release and the intracellular calcium transient. Neither effect of ryanodine was dependent on chloride ions, which suggests that chloride ions do not carry the ryanodine-sensitive current components. Strontium (2.7 mM replacing calcium) and caffeine (10 mM), two other treatments that interfere with sarcoplasmic reticulum function, had effects in common with ryanodine. This supports the hypothesis that the effects of ryanodine may be attributed to the inhibition of sarcoplasmic reticulum calcium release.     

The effects of sodium pump activity on the slow inward current in sheep cardiac Purkinje fibres

The effects of Na pump activity on the slow inward current, Isi, magnitude and twitch tension were investigated in sheep cardiac Purkinje fibres. A two-microelectrode voltage-clamp method was used, tension being measured simultaneously. Na pump activity was lowered either by reducing the extracellular K concentration, [K]O, or by applying the cardiotonic steroid strophanthidin. Reduction of [K]O from 4 to 0 mM leads to time-dependent increases in Isi magnitude and twitch tension. The increases of Isi and tension could be reversed by adding Tl, Rb, Cs or NH4 ions to the K-free superfusate. The actions of these ions are attributed to the known ability of these cations to activate the external site of the Na pump. This conclusion is supported by the observation that such activator cations do not reverse the increases in Isi and tension produced by strophanthidin. We conclude that the effects of low [K]O on Isi are mediated by Na pump inhibition. Similarly the Na pump inhibition produced by strophanthidin increases Isi and tension, although, in this case, other mechanisms may also contribute. Measurements of the activity of the electrogenic Na pump show that elevated intracellular Na ion concentration secondary to Na pump inhibition and not the instantaneous Na pump turnover rate mediates the increase in Isi magnitude.     

Relationships between voltage and tension in sheep cardiac Purkinje fibers

The two-microelectrode technique of voltage clamping sheep cardiac Purkinje fibers was used to examine the changes in contraction which occur during trains of voltage clamps. (A "train" is defined as a series of voltage clamps delivered at a particular rate, beginning after a rest long enough that the effects of previous stimulation have died away.) Contractions showed striking staircases, or progressive changes in peak isometric tension, during trains. Short clamps, clamps to voltages more negative than --20 or --30 mV, or holding potentials less negative than the resting potential favored negative staircases, while long clamps, clamps to positive voltages, and holding potentials near the resting potential each favored positive staircases. The staircase behavior appeared to be due to changes in the initial rate of recovery of the ability to contract. The changes in staircase behavior as a function of clamp voltage suggested that the relationship between peak tension and clamp voltage should depend on the experimental design. When the steady-state contraction was plotted as a function of clamp voltage, voltage-tension relations like those recently reported for working ventricle were obtained, with a threshold between --30 and - -40 mV and a steep relation between tension and voltage. When the first contraction after a rest was plotted, the threshold voltage was more negative, the curve was flatter, and the peak tensions at inside positive voltages were reduced.