An analysis of the mammalian ventricular action potential

Summary A Hodgkin-Huxley model for ventricular excitation is abstracted from electrophysiological data. A singular perturbation analysis of the 8-dimensional phase portrait of the model characterizes the role of calcium during the plateau phase of the ventricular action potential and demonstrates how the calcium refractory period prevents tetanization. Supported in part by the Undergraduate Research Opportunities Program, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA     

Theoretical simulation of the calcium action potential in squid giant synapse: Repetitive stimulation

A biophysical model of the experimentally observed calcium action potential (CAP) in squid giant synapse is proposed. Whereas the inclusion of the inward calcium current in the Hodgkin-Huxley model can generate the rising phase of CAP, to account for the observed termination of the action potential, a repolarizing process needs to be introduced. Adding a term representing Ca-activated K current, the observed features of CAP can be reproduced. However, one feature of CAP, namely the gradual shortening of the plateau duration on repetitive stimulation, cannot be simulated by this model. In this paper, it is proved that both the termination of the action potential and the gradual shortening of the plateau cannot be accounted for by inclusion of a single repolarizing process. One more repolarizing process, namely a slow voltage-dependent Ca-inactivation, is therefore proposed to account for all the observed features of CAP.     

Theoretical simulation of the calcium action potential in squid giant synapse: The plateau termination

A biophysical model is proposed for the simulation of the experimentally observed calcium action potential at the squid giant synapse. It is observed that while Ca activation at the synapse is responsible for the generation of the upstroke of the action potential, a repolarizing process needs to be invoked to simulate the plateau termination and other long-time effects. Out of the likely candidates, the Ca-activated K current has been chosen as the most plausible repolarizing process. The model can reproduce all the observed features of calcium action potential excepting its behaviour after repetitive stimulation.     

The rate of action of tetrodotoxin on sodium conductance in the squid giant axon.

When tetrodotoxin is applied to or washed away from the squid giant axon, the rates at which the sodium conductatnce is blocked and unblocked are an order of magnitude smaller than those reported for the isolated node of Ranvier. This slowing is to be expected if in squid the tetrodotoxin binding sites act as a saturable sink in series with the barrier to free diffusion imposed by the presence of the Schwann cell. A comparison has been made between the rates observed experimentally and those calculated for a computer model of the system, in order to estimate the apparent density in the membrane of both specific and non-specific tetrodotoxin binding sites. The figure thus obtained for the number of sodium channels in the squid giant axon, several hundred per square micrometre, agrees well with those derived from other lines of argument.     

The effect of displacement current on the measurement of reversal potential in squid giant axon

The measurement of the sodium reversal potential (Erev), as that potential where the early current reverses during voltage clamp, was found to exceed the true Erev by 4.1 +/- 2.4 mV (mean +/- SD) in squid giant axon. This error was found in both intact and internally perfused axons and is due to interference from the displacement current. This was shown by subtraction of the current records obtained before and after treatment with tetrodotoxin (TTX). The error in Erev is proportional to (Td/gNA)12 where Td is the time constant of the displacement current.     

The effect of temperature on veratridine action in squid giant axons

Resting membrane potential and intracellular sodium and potassium concentrations were determined at 5 and 21°C in normal and veratridine-treated axons of the squid Doryteuthis plei. 300 μM veratridine produced an increase in the intracellular sodium concentration, which changed from 52 to 284 mM in 10 min of exposure at 21°C, and from 76 to 260 mM at 5°C. Under the same treatment the intracellular potassium concentration changed from 357 to 221 mM (21°C) and from 334 to 194 mM (5°C). All the changes could be prevented by adding 1 μM tetrodotoxin. Veratridine (30, 100 and 300 μM) increased the resting sodium permeability of the giant axon, and the effect was greater at 21°C. The affinity of the membrane for veratridine increases when the nerves are cooled, the three concentrations tested produce maximum activation of the sodium channels at 5°C. But only the higher two concentrations are saturating at 21°C.     

Effects of bath resistance on action potentials in the squid giant axon: myocardial implications.

This study presents a simplified version of the quasi-one-dimensional theory (Wu, J., E. A. Johnson, and J. M. Kootsey. 1996. A quasi-one-dimensional theory for anisotropic propagation of excitation in cardiac muscle. Biophys. J. 71:2427-2439) with two components of the extracellular current, along and perpendicular to the axis, and a simulation and its experimental confirmation for the giant axon of the squid. By extending the one-dimensional core conductor cable equations, this theory predicts, as confirmed by the experiment, that the shapes of the intracellular and the extracellular action potentials are related to the resistance of the bath. Such a result was previously only expected by the field theories. The correlation between the shapes of the intracellular and the extracellular potentials of the giant axon of the squid resembles that observed during the anisotropic propagation of excitation in cardiac muscle. Therefore, this study not only develops a quasi-one-dimensional theory for a squid axon, but also provides one possible factor contributing to the anisotropic propagation of action potentials in cardiac muscle.     

Effects of fatty acids on membrane currents in the squid giant axon

Summary The effects of fatty acids on the ionic currents of the voltage-clamped squid giant axon were investigated using intracellular and extracellular application of the test substances. Fatty acids mainly suppress the Na current but have little effect on the K current. These effects are completely reversed after washing with control solution. The concentrations required to suppress the peak inward current by 50% and Hill number were determined for each fatty acid. ED₅₀ decreased about 1/3 for each increase of one carbon atom. The standard free energy was –3.05 kJ mole^–1 for CH₂. The Hill number was 1.58 for 2-decenoic acid. The suppression effect of the fatty acids depends on the number of carbon atoms in the compounds and their chemical structure. Suppression of the Na current was clearly observed when the number of carbon atoms exceeded eight. When fatty acids of the same chain length were compared, 2-decenoic acid had strong inhibitory activity, but sebacic acid had no effect at all on the Na channel. The currents were fitted to equations similar to those proposed by Hodgkin and Huxley (J. Physiol. (London) 117:500–544, 1952) and the changes in the parameters of these equations in the presence of fatty acids were calculated. The curve of the steady-state activation parameter (m ) for the Na current against membrane potential and the time constant of activation ({ie113-1}) were shifted 20 mV in a depolarizing direction by the application of fatty acids. The time constant for inactivation ({ie113-2}) was almost no change by application of the fatty acids. The time constant for activation ({ie113-3}) of K current was shifted 20 mV in a depolarizing direction by the application of the fatty acids.     

Phosphorylation of K+ channels in the squid giant axon. A mechanistic analysis

Protein phosphorylation is an important mechanism in the modulation of voltage-dependent ionic channels. In squid giant axons, the potassium delayed rectifier channel is modulated by an ATP-mediated phosphorylation mechanism, producing important changes in amplitude and kinetics of the outward current. The characteristics and biophysical basis for the phosphorylation effects have been extensively studied in this preparation using macroscopic, single-channel and gating current experiments. Phosphorylation produces a shift in the voltage dependence of all voltage-dependent parameters including open probability, slow inactivation, first latency, and gating charge transferred. The locus of the effect seems to be located in a fast 20 pS channel, with characteristics of delayed rectifier, but at least another channel is phosphorylated under our experimental conditions. These results are interpreted quantitatively with a mechanistic model that explains all the data. In this model the shift in voltage dependence is produced by electrostatic interactions between the transferred phosphate and the voltage sensor of the channel.     

Molecular motion underlying activation and inactivation of sodium channels in squid giant axons

Summary Measurements of the changes in birefringence associated with changes in membrane potential were made with internally perfused squid giant axons in low sodium solutions at 0–8°C. The time course of the birefringence changes share many properties of the gating (polarization) currents previously studied in this nerve. Both can be demonstrated as an asymmetry in the response to voltage pulses symmetrical about the resting potential which is not present about a hyperpolarized holding potential. Both have a rapid relaxation, which precedes the sodium permeability change. Both exhibit an initial delay or rising phase. Both are reversibly blocked by perfusion with 30mm colchicine; neither are altered by changes on sodium concentrations or 300nm tetrodotoxin. The birefringence response has a decrease in the amplitude of the rapid relaxation associated with the appearance of a slow relaxation. This is similar to the immobilization of fast gating charges which parallels sodium current inactivation.The amplitude of the birefringence and the gating current responses is consistent with a change in the alignment of several hundred peptide bonds per sodium channel.     

镉对豚鼠乳头肌动作电位的影响

目的:研究氯化镉(CdCl2)对正常豚鼠心室乳头肌细胞动作电位(AP)的影响。方法:用常规细胞内微电极方法记录乳头肌细胞AP。结果:在生理条件下,CdCl2可使APo期振幅(APA)和最大除极速率(Vmax)降低,动作电位时程(APD)缩短,且具有剂量依赖性。结论:CdCl2可使心室肌AP的APA、Vmax、APD发生改变,提示CdCl2有抑制Na 、Ca2 内流和激活K 外流作用。     

On the relation between displacement currents and activation of the sodium conductance in the squid giant axon.

The early time course of the current passing across the membrane in squid giant axons in which the ionic currents have been blocked reveals substantial asymmetries during and after the application of hyperpolarizing and depolarizing voltage-clamp pulses of identical size. Since the integral of the 'on' and 'off' current transients is zero, these currents must result from charge movements confined to the membrane and, therefore, they are nonlinear displacement currents. The steady state rearrangement of the charges as a consequence of sudden displacements of the membrane potential is consistent with a Boltzmann distribution of charges between two states characterized by different energy levels. Following changes in membrane potential the charges undergo a first order transition between these states. The relaxation time constant for the transition at a given temperature is a function of membrane potential. We propose that these displacement currents arise from a redistribution of the charges involved in the sodium gating system.     

雌、孕激素联合应用对豚鼠心室乳头肌细胞动作电位及收缩力的影响

目的:观察不同浓度17β雌二醇(E2)与孕酮(P)联合运用时豚鼠心室乳头肌细胞动作电位及心肌收缩力的影响.方法:采用常规玻璃微电极技术记录豚鼠心室乳头肌细胞动作电位,生理二道记录仪与示波器连接记录收缩力.结果:①E2与P单独作用可使动作电位时程(APD90、APD)延长,APD20缩短,心肌收缩力降低,APA、Vmax降低.②不同浓度E2与P联合应用,P可加强E2的效应.结论:E2与P可能抑制Ca2 通道、K 通道、Na 通道,不同浓度E2与P联合应用,P可加强E2的效应,这种作用可能呈现浓度依赖性,提示在临床应用中加用孕激素应以低浓度为益.     

葛根素对大鼠心室肌细胞动作电位及钾通道电流的影响

目的:观察葛根素对大鼠心室肌细胞动作电位及钾通道电流的影响。方法:用常规微电极方法记录大鼠心室肌细胞动作电位,用全细胞膜片钳技术记录游离心室肌细胞钾离子流。结果:不同浓度的葛根素均能延长大鼠心室肌细胞动作电位时程(APD)及抑制内向整流钾电流,具有明显的浓度依赖关系。结论:葛根素延长APD,抑制内向整流钾电流,可能是其抗心律失常的机制。     

The relationship between the inactivating fraction of the asymmetry current and gating of the sodium channel in the squid giant axon

A quantitative comparison between the voltage dependence of the inactivating component of the asymmetrical charge transfer in the squid giant axon and that of the sodium conductance indicates that activation of the sodium system involves either three subunits operating in parallel or a three-step series mechanism. This is confirmed by an examination of the relative timing of the flow of asymmetry and ionic currents during the opening and closing of the sodium channels. In agreement with previous suggestions, inactivation is coupled sequentially to activation. The evidence appears to argue against a trimeric system with three wholly independent subunits and favours a monomeric system that undergoes a complex sequence of conformational changes.     

乙醇对大鼠心肌动作电位及人Kv1.5通道的影响

为了研究乙醇对心肌动作电位的作用及其机制,本实验采用标准玻璃微电极细胞内记录技术记录离体大鼠心肌细胞的动作电位(action potential,AP),采用全细胞膜片钳技术记录HEK293细胞上表达的人Kv1.5(human Kv1.5,hKv1.5)通道电流,观察6.25、12.5、25.0、50.0、100.0及...     

Effects of arachidonic acid and the other long-chain fatty acids on the membrane currents in the squid giant axon

Summary The effects of arachidonic acid and some other long-chain fatty acids on the ionic currents of the voltage-clamped squid giant axon were investigated using intracellular application of the test substances. The effects of these acids, which are usually insoluble in solution, were examined by using -cyclodextrin as a solvent. -cyclodextrin itself had no effect on the excitable membrane. Arachidonic acid mainly suppresses the Na current but has little effect on the K current. These effects are completely reversed after washing with control solution. The concentration required to suppress the peak inward current by 50% (ED₅₀) was 0.18mm, which was 10 times larger than that of medium-chain fatty acids like 2-decenoic acid. The Hill number was 1.5 for arachidonic acid, which is almost the same value as for medium-chain fatty acids. This means that the mechanisms of the inhibition are similar in both long- and medium-chain fatty acids. When the long-chain fatty acids were compared, the efficacy of suppression of Na current was about the same value for arachidonic acid, docosatetraenoic acid and docosahexaenoic acid. The suppression effects of linoleic acid and linolenic acid on Na currents were one-third of that of arachidonic acid. Oleic acid had a small suppression effect and stearic acid had almost no effect on the Na current. The currents were fitted to equations similar to those proposed by Hodgkin and Huxley (Hodgkin, A.L., Huxley, A.F. (1952)J. Physiol (London) 117:500–544) and the change in the parameters of these equations in the presence of fatty acids were calculated. The curve of the steady-state activation parameter (m ) for the Na current against membrane potential and the time constant of activation ( _m ) were shifted 10 mV in a depolarizing direction by the application of fatty acids. The time constant for inactivation ( _h ) has almost unaffected by application of these fatty acids.