Hypothermia may induce hypokalemia and increase intracellular Ca2+ by affecting serum K+ and Ca2+ fluxes across the cell membrane. These ionic alterations may significantly change the electrophysiologic characteristics of the cardiac action potential and may induce cardiac arrhythmias. The current study was undertaken to determine whether electrophysiologic changes in Purkinje fibers induced by hypothermia could be reversed by manipulating the extracellular K+ and transmembrane Ca2+ fluxes by Ca2+ channel blockade with verapamil.


A conventional microelectrode method was used to determine the effects of hypothermia (32 +/- 0.5 degrees C and 28 +/- 0.5 degrees C) and various external K+ concentrations ([K+]o) (2.3, 3.8, and 6.8 mM) on maximum diastolic potential, maximum rate of phase 0 depolarization (Vmax), and action potential duration (APD) at 50% (APD50) and at 95% (APD95) repolarization in isolated canine cardiac Purkinje fibers. To evaluate the contribution of the slow inward Ca2+ current to action potential changes in hypothermia, the experiments were repeated in the presence of the Ca(2+)-channel antagonist verapamil (1 microM).


Variations of [K+]o induced the expected shifts in maximum diastolic potential, and hypothermia (28 degrees C) induced moderate depolarization, but only when [K+]o was > or = 3.9 mM (P < 0.05). Hypothermia decreased Vmax at all [K+]o studied (P < 0.05). Regardless of the temperature, Vmax was not affected by verapamil when [K+]o was < or = 3.9 mM, but at 6.8 mM [K+]o in hypothermia Vmax was significantly lower in the presence of verapamil. Hypothermia increased both the APD50 and the APD95. The effects of verapamil on APD were temperature and [K+]o dependent; between 37 degrees C and 28 degrees C with 2.3 mM [K+]o in the superfusate, verapamil did not affect APD. At 28 degrees C in the presence of verapamil, the APD50 and APD95 decreased only if the [K+]o was > or = 3.9 mM.


Verapamil and K+ supplementation in hypothermia may exert an antiarrhythmic effect, primarily by reducing the dispersion fo prolonged APD.

Key words: Calcium channel, antagonist; verapamil. Heart, electrophysiology: action potential duration; maximum diastolic potential; maximum rate of phase 0 depolarization. Ions, potassium: extracellular. Temperature: hypothermia.

HYPOTHERMIA can occur accidentally after exposure to a cold environment or operating room or may be intentionally induced to protect tissues against hypoxia during surgery. [1] Various cardiac arrhythmias, including ventricular fibrillation and asystole, occur at temperatures below about 30 degrees Celsius. [2] An anesthesiologist may be faced with this problem in the critical care setting while treating the patient with accidental hypothermia, intraoperatively during recovery from low temperature cardioplegia, or even during irrigation of the thoracic cavity with cold solution. [4] Major electrocardiographic manifestations of hypothermia include decreased myocardial conduction velocity with increases in PR and QT intervals and QRS complex duration. [2] At least in part, these changes can be attributed to alterations in the electrophysiologic properties of the cardiac Purkinje fibers [4] or serum electrolyte changes [5] induced by the hypothermia. [6,7] In vivo, nonhomogeneous tissue cooling slows action potential propagation and can induce differences in repolarization at various sites in the ventricular conducting system. [8] This repolarization heterogeneity facilitates nonuniform distribution of impulse conduction throughout the myocardium and may result in reentrant arrhythmias. [8–11] In addition, low temperature modifies cation conductance across cell membranes [12] and may result in clinically significant hypokalemia, which can be attributed to a temperature-mediated redistribution of Potassium sup + among body compartments. [6,7] The resting membrane potential of cardiac Purkinje fibers may depolarize in deep hypothermia, [13] and an increase in the action potential duration (APD) is seen even during mild hypothermia and hypokalemia. [14] These electrophysiologic changes can depress impulse conduction and may induce reentry arrhythmias, abnormal forms of automaticity, and heart block. [5].

The exact mechanism by which hypothermia increases cardiac irritability is unknown, but the hypothermia-induced electrophysiologic changes in Purkinje fiber action potentials may be involved. In addition to affecting Potassium sup + homeostasis. [6,7] hypothermia also increases intracellular myocardial Calcium2+ concentration ([Calcium2+]i). [15] Inhibition of the Sodium sup +-Potassium sup + pump activity by low temperature leads to an increase in intracellular Sodium sup + concentration ([Sodium sup +]i), which may promote Calcium2+ influx during the plateau of the action potential or oppose a Calcium2+ efflux during diastole by the Sodium sup +-Calcium2+ exchange mechanism. [16] The excessive accumulation of intracellular Calcium2+, termed “Calcium2+ overload,”[17] may result in delayed afterdepolarizations, causing severe cardiac arrhythmias. [5,17–19] Furthermore, it has been suggested that hypothermia, by delaying inactivation of the inward Calcium2+ current (ICa) and maintaining the ICafor a longer time, contributes to action potential lengthening and arrhythmias based on abnormal impulse propagation. [20].

We hypothesized that if increased [Calcium2+]iunderlay important changes in electrophysiologic characteristics of Purkinje fibers in hypothermia, especially prolongation of the APD, we should be able to alter their course with a Calcium2+-channel blocking agent. Because hypothermia alters Potassium sup + homeostasis, [6,7] we also examined the effects of hypothermia on action potential characteristics over the wide range of external [Potassium sup +]([Potassium sup +]o) in the absence and presence of verapamil. The reduction of APD with verapamil, if present, might be expected to reduce the incidence of hypothermia-induced arrhythmias, specifically dysrhythmias based on regional differences in myocardial repolarization.

This study was approved by the Medical College of Wisconsin Animal Care Committee and conformed with standards set forth in the Guide for Care and Use of Laboratory Animals.*

Adult mongrel dogs (10–22 kg, of either sex)(n = 45) were anesthetized with 30 mg/kg pentobarbital sodium. The heart was quickly excised, and the anterior false tendon with attached papillary muscle from the left ventricle was removed and immersed in modified Krebs' solution (22 degrees Celsius) equilibrated with 97% Oxygen2and 3% CO2. Small (< 1-cm2) preparations with free-running strands of Purkinje fibers were dissected from this tissue and pinned to the silicone elastomere floor of a 2-ml chamber and superfused at a rate of 4 ml/min with modified Krebs' solution (37 degrees Celsius) containing 2.3, 3.9, or 6.8 mM KCl with or without 1 micro Meter verapamil and equilibrated with a 97% Oxygen2-3% CO2gas mixture. The millimolar composition of Krebs' solution was 137 NaCl, 12 NaHCO3, 1.8 NaH2PO4, 1.8 CaCl2, 0.5 MgCl2, 5.5 glucose, and 0.05 EDTA, with a pH of 7.4.

Each preparation was stimulated at a constant rate (1 Hz) with the use of bipolar-silver wire endocardial surface electrodes. The stimuli were square-wave pulses lasting 2 ms at 1.5 times threshold. Transmembrane action potentials were recorded with conventional microelectrode techniques. Action potential changes stabilized within 5–8 min. Glass microelectrodes (15–30-M Omega resistance) were coupled by Silver-AgCl wire to a preamplifier (World Precision Instruments, New Haven, CT). Action potential signals were recorded on frequency-modulated tape (AR Vetter, Rebersburg, PA) for later analysis of maximum diastolic potential (MDP), maximum rate of phase 0 depolarization (Vmax), and APD at 50%(APD50) and at 95%(APD sub 95) repolarization. These values were displayed and measured electronically directly off the digital oscilloscope (Nicolet 310). The Vmaxwas determined with a differentiator exhihiting a linear response from 100–1,000 V/s. The “zero” potential was obtained at the beginning and at the end of the experiments by withdrawing the microelectrode from the inside of the fiber. Because the ground connection between the bath and the circuit was made through a direct Silver-AgCl connection, the change in temperature may slightly influence the half cell potential in the bath, and this change will sum with the real MDP changes. To verify the significance of this phenomenon we performed additional experiments (n = 8) in which the microelectrode was placed into the bath, the temperature was changed from 37 degrees Celsius to 28 degrees Celsius, and the Silver-AgCl bath junction potential change was measured. The half cell potential caused a hyperpolarization to be measured by the microelectrode over this temperature range by -2.1 plus/minus 0.02 mV and -4.2 plus/minus 0.05 mV at 32 degrees Celsius and 28 degrees Celsius respectively. All measured MDP values at 32 degrees Celsius and 28 degrees Celsius were corrected for the above differences, by subtracting the temperature-induced bath potential from the actual measured potential, and all the statistical analyses were performed with these corrected values.

The tissue bath was surrounded by a thermostatically controlled water bath, maintained at a constant temperature of 37 plus/minus 0.02 degrees Celsius. The low bath superfusate temperatures (32 and 28 plus/minus 0.5 degrees Celsius) were attained by readjusting the setting of the thermostat. The temperature in the tissue bath was gradually decreased from 37 degrees Celsius to 25 degrees Celsius over a 20–30-min period. The temperature of the solution was measured by a small, rapidly responding, custom made thermistor probe placed less than 2 mm from the preparation. The fluid level in the chamber was kept at a constant height (4 mm) by continuous suction.

The preparations were allowed to equilibrate for about 1 h. To determine the effects of hypothermia on action potential characteristics, action potentials were recorded at 37 degrees Celsius, 32 degrees Celsius, and 28 degrees Celsius (plus/minus 0.5 degrees Celsius). To determine the effects of various [Potassium sup +]oon action potentials, experiments were performed with 2.3, 3.9, and 6.8 mM [Potassium sup +] in the superfusate. Verapamil (Sigma, St. Louis, MO) prepared as stock solution (100 micro Meter) was added to measured volumes to achieve the desired 1 micro Meter concentration in the superfusate. The same set of action potential measurements were performed with 1 micro Meter verapamil in the superfusate. Tissues were exposed to each [Potassium sup +]oand to verapamil (1 micro Meter) for 20 min before measurement of action potential characteristics.

Data are expressed as means plus/minus standard error of the mean. Statistical analysis was performed by paired and unpaired t tests and with one-way analysis of variance (analysis of variance repeated measures and factorial analysis), as appropriate, with P < 0.05 considered statistically significant.

Typical effects of hypothermia on the action potential in canine Purkinje fibers are illustrated in the upper panel of Figure 1. Each preparation was stimulated at a constant rate of 1 Hz. Recordings were taken from the same cell at various temperatures at a [Potassium sup +]oof 3.9 mM. Hypothermia significantly increased APD. The bottom panel of Figure 1shows the changes of action potential in hypothermia in the presence of 1 micro Meter verapamil in the superfusate. In the presence of verapamil the Purkinje fiber's action potential is shorter than at the same temperature without the drug (dashed action potential was recorded at 37 degrees Celsius in the absence of verapamil). The slope of phase 2 repolarization is increased by verapamil, a finding that is consistent with the blockade of a slow inward current such as that carried by a Calcium2+ ion. Hypothermia, on the other hand, appears to decrease the slope of phase 2, suggesting that low temperature affects the Calcium2+-dependent ionic mechanisms in opposite direction.

Maximum Diastolic Potential

The effects of two stages of hypothermia (32 degrees Celsius and 28 degrees Celsius) on the MDPs of Purkinje fibers were recorded during the exposure to low (2.3 mM), normal (3.9 mM), and high (6.8 mM)[Potassium sup +]oin the superfusate (Figure 2). MDPs were higher (hyperpolarized) at low, and lower (depolarized) at high [Potassium sup +]othan at normal [Potassium sup +]o(P < 0.05), regardless of the temperature. During the 20–30 min of gradual cooling from 37 degrees Celsius to 28 degrees Celsius, MDP decreased at normal and high [Potassium sup +]o(P < 0.05), and did not change at low [K sup +]o.

At 37 degrees Celsius and in the presence of verapamil MDP was lower (difference not significant, P > 0.05 vs. control;Figure 2). In hypothermia and at [K sup +]ogreater or equal to 3.9 mM loss of membrane potential was parallel to this of control Purkinje fibers, and reached significant depolarization at 28 degrees Celsius (P < 0.05). At [K sup +]o2.3 mM there was no effect of temperature on MDP in verapamil-superfused Purkinje fibers.

Maximum Rate of Phase 0 Depolarization

The effects of hypothermia, Potassium sup + variation, and verapamil on Vmaxare summarized in Figure 3. Hypothermia decreased Vmaxat each [K sup +]o(P < 0.05). Despite significant differences in MDP at various [K sup +]oat 37 degrees Celsius (Figure 2), the respective Vmaxvalues were not different (Figure 3), although there were trends for Vmaxto be lower at 6.8 and higher at 2.3 mM [K sup +]othan at 3.9 mM [K sup +]o(P > 0.05). At [K sup +]oless or equal to 3.9 mM with verapamil in the superfusate no additional effect, besides that of temperature, was noted on Vmax(Figure 3). At 6.8 mM [K sup +]oin hypothermia, Vmaxwas lower in the presence of verapamil (P < 0.05).

To quantify the effects of changing MDP on Vmax, we examined eight additional Purkinje fiber preparations by gradually increasing [K sup +]oin the superfusate from 0.8 to 10 mM; MDP and Vmaxvalues were measured at 37 degrees Celsius and at 30 degrees Celsius (Figure 4). At both temperatures MDP decreased (less negative) directly with increasing [K sup +]obetween 2.3 and 10 mM (P < 0.05); below 2.3 mM MDP did not further change (P > 0.05). Only between 3.9 and 8.5 mM [K sup +]oMDP was lower at 30 degrees Celsius (P < 0.05), Vmaxdecreased moderately between 2.3 and 6.8 mM [K sup +] sub o, followed by a rapid decrease when [K sup +]oincreased above 6.8 mM (MDP less or equal to -80 mV)(Figure 4). Compared with 2.3 mM [K sup +]o, at 0.8 mM [K sup +]othere was a significant decrease in Vmaxat 37 degrees Celsius, and no change in Vmaxat 30 degrees Celsius. By varying the [K sup +]o, Vmaxat 37 degrees Celsius and 30 degrees Celsius resulted in similar pattern of changes, yet amplitudes were different.

Action Potential Duration

Hypothermia increased early (APD50) and late (APD95) stages of repolarization (Table 1and Table 2). Figure 5and Figure 6compare the relative changes in APD50and APD95at various temperatures and [K sup +]owith and without verapamil in the superfusate. In hypothermia the increase in APD50(at 28 degrees Celsius) and APD95(at 32 degrees Celsius and 28 degrees Celsius) was inversely related to [K sup +]o; at 6.8 mM [K sup +] sub o in hypothermia APD50and APD95were always closest to normothermic control. Regardless of temperature, verapamil did not affect APD50or APD95at 2.3 mM [K sup +]o. At 28 degrees Celsius and [K sup +]ogreater or equal 3.9 mM verapamil significantly shortened the APD50(Figure 5) and the APD95(Figure 6). At 28 degrees Celsius and 6.8 mM [K sup +]owith verapamil, the APD50was 21% shorter whereas the APD95was 12% longer than normothermic control. Without the verapamil at 28 degrees Celsius, there were no longer relative differences between the increases in APD50and APD95, regardless of the [K sup +]o, indicating that hypothermia prolonged early and late repolarization similarly (Figure 7). After verapamil was introduced to the superfusate, the relative changes in APD became dependent on the actual [K sup +]o; at 2.3 mM [K sup +] APD50and APD95were unchanged (Figure 7(A)); at 3.9 mM [K sup +]othe primary effect was on APD50(Figure 7(B)); at 6.8 mM APD50and APD95decreased, with APD50> APD95(P < 0.05)(Figure 7(C)).

Induced hypothermia is commonly used to protect myocardium and other tissues against ischemia during cardiopulmonary bypass. [1] At the same time hypothermia is known for its arrhythmogenic potential, including ventricular fibrillation. [1–3] The common mechanisms underlying cardiac arrhythmias are the loss of the MDP in Purkinje fibers, [5] diastolic Calcium2+ overload, [16] and altered dispersion of repolarization, [11] all being reported in hypothermia. [4,13,14,20,21] Hypokalemia has been encountered during hypothermia, [6,7] and rebound hyperkalemia has occurred during rewarming. [7] This study examined electrophysiologic alterations of MDP, Vmax, and APD during hypothermia and hypothermic hypokalemia and their reversibility after verapamil treatment and Potassium sup + supplementation.

Maximum Diastolic Potential

In the current study, the shift in MDP caused by variation of [K sup +]obetween 2.3 and 10 mM was consistent with values predicted by the Nernst equation [14]: depolarization was present at high, and hyperpolarization at low [K sup +]o(Figure 2and Figure 4). Below [K2+]o2.3 mM, specifically at 0.8 mM, MDP did not further hyperpolarize, but also did not depolarize as could be expected. [14] We do not have a plausible explanation for the disparity between ours and previously reported findings. [14].

We found moderate loss of membrane potential (depolarization) with reduction of temperature to 28 degrees Celsius but only at [K sup +]ogreater or equal to 3.9 mM. A very small decrease in the resting membrane potential of canine Purkinje fibers was described at 25 degrees Celsius, [13] but this decrease was considerably larger below 20 degrees Celsius. [4] The decreases in MDP in hypothermia have been attributed to inhibition of adenosine triphosphate-dependent Sodium sup +-Potassium sup + pump. [22–24] In our study the Calcium24-channel blockade with 1 micro Meter verapamil had no significant additional effect on MDP regardless of the temperature, and when [K sup +]owas greater or equal to 3.9 mM moderate depolarization at 28 degrees Celsius was parallel to that of control Purkinje fibers. This tendency toward depolarization when [Potassium sup +]owas greater or equal to 3.9 mM may not be incidental, because verapamil is known to decrease MDP at higher concentrations. [25,26] Dersham and Han described a nonsignificant decrease in MDP after 1 micro Meter verapamil, and a statistically significant decrease (from 86 to 80 mM) after 2 micro Meter. [9] Similar results after 2 micro Meter of verapamil were reported by Amerini et al. [19] The mechanism responsible for MDP decrease after verapamil was attributed to the reduced Potassium sup + conductance, [19,27] but we cannot confirm this mechanism because the increase in K sup + conductance by increasing [Potassium sup +][8] in our study did not reduce the amount of depolarization induced by verapamil. It is unlikely that small changes in MDP, present at normal serum [Potassium sup +]([nearly equal] 3.9 mM), have any clinical significance. Our findings suggest that hypothermic cardiac arrhythmias, which occur around 28 degrees Celsius, cannot be attributed to hypothermia-induced membrane depolarization, providing serum Potassium sup + is in normal range.

Maximum Rate of Phase O Depolarization

The magnitude of Vmaxis determined by the fast inward Sodium sup + current (INa), and has traditionally been used as an index of Sodium sup + channel availability. [28,29] In our experiments, hypothermia decreased Vmax(Figure 3), which may be explained by the temperature-mediated decrease in INa. [21,30] Hypothermia decreases Sodium sup +-Potassium sup + adenosine triphosphatase activity, which then results in increased [Sodium sup +] sub j and decrease in driving force for INa. [31–33] At [Potassium sup +]oless or equal to 3.9 mM in the normothermic and hypothermic states, Calcium2+-channel blockade with 1 micro Meter verapamil did not alter Vmax. Although verapamil in lower concentrations has no significant effect on Vmax, [9,25,34,35] the Calcium2+-channel blockade may decrease Vmaxwhen given in higher concentrations. [30,37] However, at 6.8 mM [Potassium sup +]oin hypothermia, we found that verapamil decreased Vmax(P < 0.05)(Figure 3). It is possible that the cumulative effect of hypothermic INablockade, low MDP caused by hyperkalemia, and verapamil-induced inhibition of INa, [36,37] possibly potentiated by low temperature, resulted in a reduction of Vmax.

Because Vmaxis voltage-dependent we examined the relation between MDP and Vmaxby varying the [Potassium sup +]obetween 0.8 and 10 mM. Whereas the increase in [Potassium sup +]obetween 2.3 and 10 mM caused an almost linear decrease in MDP (depolarization), at less than 2.3 mM, MDP ceased to decrease further, consistent with a decrease in resting Potassium sup + conductance at low [Potassium sup +]o. [38] The decrease in temperature from 37 degrees Celsius to 30 degrees Celsius induced a leftward shift of the V sub max curve. Although there was a decreasing trend, Vmaxdid not significantly change when [Potassium sup +]owas increased from 2.3 to approximately 6 mM, a finding that is consistent with the results described in Figure 3. Therefore, Vmaxwas not greatly affected when the voltage (MDP) was between -100 and -85 mV (2.3–6 mM [Potassium sup +]o), but was markedly decreased when MDP became less negative than -80 mV. In extreme hypokalemia (0.8 mM) at 37 degrees Celsius Vmax, significantly decreased, at least in part mirroring changes in the respective MDP. Our data suggest that during phase 0 of the stimulated action potential at MDP between -80 and -100 mV, the fraction of available Sodium sup + channels remains relatively constant or only slightly increases as MDP becomes more negative. Thus, when MDP reaches -80 mV, the near-maximum capacity for Sodium sup + influx through the fast Sodium sup + channels may be reached (especially in hypothermia) and cannot be further increased by more negative MDP. The decrease in V sub max when MDP drops below approximately -72 mV at 37 degrees Celsius and -80 mV at 30 degrees Celsius, is very abrupt. In addition, hypothermia significantly decreased Vmaxand less affected MDP. These findings are consistent with the existence of two mechanisms that may effect Sodium sup +-channel kinetics: first, a temperature-dependent mechanism that is responsible for the decrease in Vmaxduring cooling, and second, a voltage-dependent mechanism that is active only at MDP less negative than -80 mV. In deep hypothermia and at [Potassium sup +]o6.8 mM, verapamil further decreased Vmax, most likely by the interference with INakinetics by voltage-dependent mechanism. The clinical significance of the decrease in Vmaxby verapamil and [Potassium sup +]oin hypothermia has to be studied, but typically the drugs that are able to decrease the I sub Na [39] induce the following chain of events: decrease in [Sodium sup +]i, increase in Calcium2+ efflux and therefore a decrease in sarcoplasmic reticulum Calcium2+ loading. These events may ultimately result in an antiarrhythmic effect of the drugs by decreasing oscillatory afterpotentials. [40].

Action Potential Duration

The cardiac action potential plateau is determined by a delicate balance among several distinct time- and voltage-dependent ionic currents, [8,38,41–43] After upstroke of the action potential caused by the influx of Sodium sup +, the INabecomes inactivated. This inactivation does not lead to immediate repolarization, because the initial depolarization reduces the inwardly rectifying Potassium sup + conductance and opens voltage-gated Calcium2+ channels thus permitting the intracellular influx of Calcium2+ by the INa. During the action potential plateau, while membrane conductance for all ions is reduced, several currents help to maintain the transmembrane potential at around 0 mV, including ICa, Sodium sup +“window.” Cl and inward (anomalous) rectifying Potassium sup + currents. In addition, currents produced by the electrogenic Sodium sup +-Potassium sup + pump and Sodium sup +- Calcium2+ exchange mechanism help to maintain the action potential plateau. The outward (delayed) rectifying Potassium sup + current (IK) and the inward rectifying Potassium sup + current are responsible for final rapid repolarization (phase 3). Details of the ionic basis of action potential are reviewed elsewhere. [5,16,23,27,38,41,45].

The electrophysiologic mechanisms responsible for the marked prolongation of APD seen in hypothermia are not fully understood, although mechanisms based on temperature dependence of IK[46] and ICa[47] have been proposed. One important mechanism may be the hypothermia-induced reduction of IK, which is a major contributor to membrane repolarization. [46] In addition, lengthened APD in hypothermia may be attributed to delayed inactivation of the I sub Ca sup +[47,48] which will maintain the ICafor a longer time. [20] Finally, prolonged APD may be also attributed to increase in [Calcium2+]ithrough altered Sodium sup +-Calcium2+ exchange. [33] The activity of the Sodium sup +-Potassium sup + pump is likely to be reduced in hypothermia [31,32] resulting in a net rise in [Sodium sup +]i. [33] The increase in [Sodium sup +]jmay promote Calcium2+ influx during the plateau of the action potential or reduce Calcium2+ efflux during diastole by Sodium sup +-Calcium2+ exchange. [49] Although the activity of the Sodium sup +-Calcium2+ exchanger is temperature dependent, its relatively low Q10(the rate of exchange produced by changing the temperature 10 degrees Celsius) means that this mechanism may contribute to raising diastolic free [Calcium2+]j. [44].

We have demonstrated that hypothermia equally lengthens the APD50and APD95, indicating that low temperature similarly affects the ionic mechanisms that determine the early and late stages of repolarization (Table 1and Table 2and Figure 7), i.e. earlier discussed IKand ICa. The finding that hypokalemia, similarly to hypothermia, prolongs the APD is not surprising, because low [Potassium sup +]odecreases Potassium sup + conductance. [38] APD50and APD95were shorter at 6.8 mM [Potassium sup +]othan at either 3.9 or 2.3 mM [Potassium sup +]o(Figure 5and Figure 6) as a result of improved Potassium sup + conductance. [8,40,45,50] Interventions that affect ionic currents during the repolarization phase may selectively alter the shape and duration of the cardiac action potential. Verapamil may affect both the ICaand Potassium sup + conductance. [43,51] In our study, 1 micro Meter of verapamil significantly shortened both the APD50and the APD95, but only at 28 degrees Celsius and [Potassium sup +]ogreater or equal to 3.9 mM (Figure 5and Figure 6). At this temperature and with 6.8 mM [Potassium sup +]oand verapamil in the superfusate, the relative shortening of APD50exceeded that of APD95(Figure 7(C)). The observation that earlier phases of repolarization (APD50) appear to be more readily affected by verapamil is consistent with the primary blocking action of this agent on ICa. At the same time less affected shortening of APD95by verapamil, and its significant shortening at higher [Potassium sup +]osuggests that another mechanism is involved in the lengthening of APD at low temperature, presumably hypothermia-induced reduction of IK. [46] Also, when [Potassium sup +]owas sufficiently high at 28 degrees Celsius, both APD50and APD95were shortened, indicating not only an important role of [Potassium sup +]oin increasing Potassium sup + conductance but also an interaction between Calcium2+-channel blockade and Potassium sup + conductance. Our study indicates that hypokalemia must be corrected if the APD is to be shortened with verapamil. Not only does correction of hypokalemia affects APD through increases of Potassium sup + conductance, [38] but in addition, as Cavalie et al. have shown, ICainactivation (which is delayed by hypothermia) depends on voltage and is greater at a more depolarized potential, [52] as in our study at 6.8 mM.

Different arrhythmias may arise if the [Calcium2+]iis increased or if cardiac action potential is lengthened. First, increase in [Calcium2+]icaused by hypothermia results in oscillatory release of [Calcium2+] from sarcoplasmic reticulum; this may generate the transient inward current allowing more Sodium sup + and Calcium2+ into the cell creating delayed afterdepolarization. [5,17,38] When delayed after-depolarization reaches certain threshold, arrhythmias may be triggered. These arrhythmias can be seen at low [Potassium sup +]o, and when Sodium sup + extrusion from the cell is reduced, [8] because both may be encountered during hypothermia. By diminishing [Calcium2+]iinflux, verapamil reduces Calcium2+ overload, delays afterdepolarization amplitude and reduces triggered automaticity. [53,54] Verapamil thus may be useful for treatment of hypothermia-induced arrhythmias based on delayed afterdepolarization. Second, reentry arrhythmias may occur during the propagation of slow action potentials through Purkinje fibers [5,8,21,31,55] especially in vivo, when, because of uneven tissue cooling, heterogeneity of repolarizations may ensue. [8] A shortening of APD with verapamil in hypothermia is both temperature and [Potassium sup +]odependent, and this may have significant clinical implications, but only at higher [Potassium sup +]o: the lower the temperature, the longer the APD, and the greater the shortening effect of verapamil. Because the APD is very dependent on regional myocardial temperature, verapamil selectively affects APD: a greater APD in a cooler region will be decreased more than a lesser APD in a warmer region. By reducing the regional differences in myocardial repolarization this effect may decrease the propensity for development of arrhythmogenic reentry circuits.

In conclusion, there appears to be an increased relative I sub Ca at low temperature, as evidenced by the effective shortening of APD in deep hypothermia by verapamil. Although this shortening may result from an increase in ICa, a decrease in Potassium sup + conductance at low temperature is more likely, as evidenced by the effective shortening of APD when the Potassium sup + conductance is increased by increasing [Potassium sup +]O. Finally, the effectiveness of verapamil to shorten APD in hypothermia was dependent on [Potassium sup +]Osuggesting that the increase in Potassium sup + conductance is an important factor for achieving this verapamil effect. Verapamil and Potassium sup + supplementation in hypothermia may have an antiarrhythmic effect primarily by reducing the dispersion of prolonged APD. Further studies will be needed to evaluate this antiarrhythmic effect in vivo.

The authors thank Edith Sulzer for her help in the preparation of this manuscript.

* Guide for Care and Use of Laboratory Animals. Publication 85–23. Bethesda, MD, Public Health Services, National Institutes of Health, revised 1985.

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