Time Course Of Action Potential Research Articles

Excitable Membrane, Ion Channel, Electrostatics: A Historical Sketch

The roughly one-tenth of a volt across an excitable membrane may not seem large, but across a membrane two lipid molecules thick it yields an electric field comparable to one causing atmospheric lightning. Early attempts to explain the physical basis of the nerve impulse modeled its ion currents as stemming from two processes: the movement of ionic charges by the electric field and the ion diffusion due to ion-concentration differences across the membrane. Hampered by its assumptions of constant properties, this classical electrodiffusion model couldn’t explain axonal membrane data. An eclectic approach combining an equation from electrodiffusion theory with elements of cable theory, wave theory and circuit theory—capacitor, batteries, variable conductors—transformed biophysical science with its successful fitting of the action potential's time course. The development of glass pipette electrodes that restrict ion flow to microscopic membrane patches led to the isolation of ion channels, glycoprotein macromolecules that selectively conduct ion currents. Because it's easy to visualize water-filled pores across these macromolecules, fitted with gates that block or allow ion currents across the membrane, they were named voltage-gated ion channels. This approach is challenged because of its: simplistic models of everyday devices, including screws and paddles; assumption of constant channel capacitance; and disregard of electrostatic repulsions between same-charge residues. Genetic engineering techniques brought the discovery of structural features of many such ion channels. One universal feature is a membrane-spanning segment with a regular array of positively charged amino-acid residues. All also contain multiple amino acids whose sidechains fork into two branches. An electrostatic model based on the ion channel's strong similarities to ferroelectric liquid crystals containing branched chains helps explain the depolarization-driven activation of voltage-sensitive ion channels. [See Leuchtag H R (2008) Voltage-Sensitive Ion Channels, Springer; (2017) Biophys. J. 112(3):544a; 112(3):543a-544a; 112(3):464a.]

Computational analysis of the electromechanical consequences of short QT syndrome.

Computational analysis of the electromechanical consequences of short QT syndrome.

Rapid time course of action potentials in spines and remote dendrites of mouse visual cortex neurons

Axonally initiated action potentials back-propagate into spiny dendrites of central mammalian neurons and thereby regulate plasticity at excitatory synapses on individual spines as well as linear and supralinear integration of synaptic inputs along dendritic branches. Thus, the electrical behaviour of individual dendritic spines and terminal dendritic branches is critical for the integrative function of nerve cells. The actual dynamics of action potentials in spines and terminal branches, however, are not entirely clear, mostly because electrode recording from such small structures is not feasible. Additionally, the available membrane potential imaging techniques are limited in their sensitivity and require substantial signal averaging for the detection of electrical events at the spatial scale of individual spines. We made a critical improvement in the voltage-sensitive dye imaging technique to achieve multisite recordings of backpropagating action potentials from individual dendritic spines at a high frame rate. With this approach, we obtained direct evidence that in layer 5 pyramidal neurons from the visual cortex of juvenile mice, the rapid time course of somatic action potentials is preserved throughout all cellular compartments, including dendritic spines and terminal branches of basal and apical dendrites. The rapid time course of the action potential in spines may be a critical determinant for the precise regulation of spike timing-dependent synaptic plasticity within a narrow time window.

Do Mouse Epicardial Action Potentials Present Phase 2?

Mouse ventricular action potentials (AP) recorded from acute dissociated cardiac myocytes lacks of phase 2. In order to evaluate if this is always true for all the muscular layers within the ventricular wall, we decided to perform optical and electrophysiological experiments to evaluate the time course of APs in the subepicardial layer of intact perfuse mouse hearts. Epicardial APs show a typical spike and dome morphology present in other non-rodent mammalian species. The APs recorded optically by means of Pulsed Local Field Fluorescence Microscopy and the potentiometric dye Di-8-Anneps show a very similar time course when compared to the one obtained with microelectrodes. However, the phase 2 of the optically recorded APs is more depolarized in comparison with the electrically recorded one suggesting a larger contribution of the t-system to the epicardial APs. In order to evaluate the molecular mechanisms involved in the genesis of phase 2 APs in mouse epicardium we performed experiments to evaluate the role of intracellular Ca2+ release on the time course of APs. Interestingly, both ryanodine/thapsigargin treatment and perfusion with caffeine significantly decreased the contribution of phase 2 to the APs. Finally we evaluate the hypothesis that intracellular Ca2+ is traduced in AP changes by an activation of influx of Na+ through the Na Ca exchanger in the forward mode. Experiments where extracellular Na+ was replaced by increasing concentration of Li+ induce a significant decrease of the AP phase 2. Moreover, two known blockers, KB-R7943 (KBR) and SEA0400 (SEA) also dramatically decrease the contribution of phase 2 to the AP time course. Our results indicate that mouse epicardial AP displays a significant phase 2 that is generated by an influx of Na through the Na Ca exchanger. Supported by NIH R01-HL-084487 to AE.

Quantitative classification of somatostatin-positive neocortical interneurons identifies three interneuron subtypes

Deciphering the circuitry of the neocortex requires knowledge of its components, making a systematic classification of neocortical neurons necessary. GABAergic interneurons contribute most of the morphological, electrophysiological and molecular diversity of the cortex, yet interneuron subtypes are still not well defined. To quantitatively identify classes of interneurons, 59 GFP-positive interneurons from a somatostatin-positive mouse line were characterized by whole-cell recordings and anatomical reconstructions. For each neuron, we measured a series of physiological and morphological variables and analyzed these data using unsupervised classification methods. PCA and cluster analysis of morphological variables revealed three groups of cells: one comprised of Martinotti cells, and two other groups of interneurons with short asymmetric axons targeting layers 2/3 and bending medially. PCA and cluster analysis of electrophysiological variables also revealed the existence of these three groups of neurons, particularly with respect to action potential time course. These different morphological and electrophysiological characteristics could make each of these three interneuron subtypes particularly suited for a different function within the cortical circuit.

GABAergic Signaling Increases Through the Postnatal Development to Provide the Potent Inhibitory Capability for the Maturing Demands of the Prefrontal Cortex

The developmental profile of the firing patterns and construction of synapse connection were studied in LTS interneurons of prefrontal cortex (PFC) in rats with age (from P7 to P30). We used whole cell patch-clamp recordings to characterize electrophysiological properties of LTS interneurons in PFC at different age stages, including the action potentials (APs), short-term plasticity (STP), evoked excitatory postsynaptic currents (eEPSCs), spontaneous excitatory postsynaptic currents (sEPSC), and spontaneous inhibitory postsynaptic current (sIPSC). The developmental profile of LTS interneurons in our research showed two phases changes. The early phase from P7-P11 to P16-P19 during which the development of individual LTS interneuron dominated and just some simple synaptic connections formed, the synaptic inputs from pyramidal cells play a promoting role for the maturation of LTS interneurons to some extent. This was based on the changes of APs, eEPSCs, and STP such as the curtailment of time course of APs, the increasing facilitation of STP before P16-P19 group. The late phase from P20-P23 to P > 27 during which the function of inhibitory cortex network enhanced and the characters of this inhibitory cortex network continually changed although in the oldest age group (P > 27) in our research. The frequency and amplitude of sIPSC showed continually changes, and at the same age group, the frequency ratios and amplitude ratios of sIPSC was higher than that of sEPSC. Our study showed a foundation to clarify mechanisms underlying the evolution in time of intrinsic neuronal membrane properties and their important roles in balancing the cortex network, providing an academic foundation for the pathological researching on some psychiatric and neurological disorders.

Targeting calcium handling in arrhythmias

Abnormal calcium (Ca) handling can contribute to arrhythmogenesis directly by triggering abnormal depolarizations and indirectly by modulating action potential time course and duration. Recent data have shown the importance of these mechanisms in rare genetic diseases but also in more common conditions such as heart failure. Modulating Ca release from the sarcoplasmic reticulum via the ryanodine receptor, Ca uptake via sarcoplasmic reticulum Ca ATPase or Ca removal from the cell via the Na/Ca exchanger, are potential approaches to reduce arrhythmias. New tools allow exploring these ideas. The principles underlying this approach and the first results are critically reviewed.

Functional Imaging, Spatial Reconstruction, and Biophysical Analysis of a Respiratory Motor Circuit IsolatedIn Vitro

We combined real-time calcium-based neural activity imaging with whole-cell patch-clamp recording techniques to map the spatial organization and analyze electrophysiological properties of respiratory neurons forming the circuit transmitting rhythmic drive from the pre-Bötzinger complex (pre-BötC) through premotoneurons to hypoglossal (XII) motoneurons. Inspiratory pre-BötC neurons, XII premotoneurons (preMNs), and XII motoneurons (MNs) were retrogradely labeled with Ca(2+)-sensitive dye in neonatal rat in vitro brainstem slices. PreMN cell bodies were arrayed dorsomedially to pre-BötC neurons with little spatial overlap; axonal projections to MNs were ipsilateral. Inspiratory MNs were distributed in dorsal and ventral subnuclei of XII. Voltage-clamp recordings revealed that two currents, persistent sodium current (NaP) and K(+)-dominated leak current (Leak), primarily contribute to preMN/MN subthreshold current-voltage relationships. NaP or Leak conductance densities in preMNs and MNs were not significantly different. We quantified preMN and MN action potential time course and spike frequency-current (f-I) relationships and found no significant differences in repetitive spiking dynamics, steady-state f-I gains, and afterpolarizing potentials. Rhythmic synaptic drive current densities were similar in preMNs and MNs. Our results indicate that, despite topographic and morphological differences, preMNs and MNs have some common intrinsic membrane, synaptic integration, and spiking properties that we postulate ensure fidelity of inspiratory drive transmission and conversion of synaptic drive into (pre)motor output. There also appears to be a common architectonic organization for some respiratory drive transmission circuits whereby many preMNs are spatially segregated from pre-BötC rhythm-generating neurons, which we hypothesize may facilitate downstream integration of convergent inputs for premotor pattern formation.

Effects of sigma receptor ligand haloperidol on ionic currents in rat cardiomyocyte

Sigma receptor ligand haloperidol is a neuroleptic drug with adverse cardiovascular effects, mainly ventricular arrhythmias. Its effects on the time course of action potential and on the basic ionic currents (sodium current INa, calcium current ICa, and potassium currents Ito,, IK1 and IK,end - current at the end of 250 ms pulse) were studied on enzymatically isolated rat ventricular cardiomyocytes using the whole cell patch clamp technique at room temperature. Haloperidol induced reversible and concentration-dependent inhibition of all investigated membrane currents. 1 ľmol/l inhibited INa by 39 %, ICa by 19.5 %, Ito by 23 % and IK,end by 14%, 10 ľmol/l haloperidol - by 95 %, 22 % 80 % 37 %, respectively, and IK1 by 29 %. Reversible loss of action potential in haloperidol was observed. Inhibition of Ito-amplitude by the drug was accompanied by acceleration of apparent inactivation. Both were voltage-independent. The time course of Ito recovery from inactivation was decelerated in the drug and 9 % of Ito-channels recovered with slow time constant about 1.4 s. It corresponded with a cummulation of inhibition at higher stimulation frequencies (3.3 Hz). We conclude that haloperidol inhibited all the main ionic current in rat cardiomyocytes with profound inhibition of INa that corresponded with loss of action potential. Inhibition of Ito is very likely caused by interaction of haloperidol with Ito-channels in open and in inactivated states as supported by the results of mathematical simulations.

Late Post-myocardial Infarction Induces a Tetrodotoxin-resistant Na+Current in Rat Cardiomyocytes

Left ventricular remodeling after myocardial infarction is accompanied by electrical abnormalities that might predispose to rhythm disturbances. To get insight into the ionic mechanisms involved, we studied myocytes isolated from four different regions of the rat ventricles, 4–6 months after ligation of the left coronary artery. Using the whole-cell patch-clamp technique, we never observed T-type Ca2+current in both diseased and control hearts. In contrast, in 41 out of 78 cells isolated from 16 post-myocardial infarcted rats, analysed in the presence of 30 m m Na+ions, we found a tetrodotoxin (TTX)-resistant Na+current with quite variable amplitude in every investigated region. Albeit being resistant to 100 μM TTX, this Na+-dependent current was highly sensitive to lidocaine since 3 μM lidocaine induced about 65% tonic block. It was also inhibited by 5 μM nifedipine and 2 m m Co2+, but was insensitive to 100 μM Ni2+. The TTX-resistant Na+channel availability was shifted rightward by 25–30 mV with respect to TTX-sensitive Na+current; therefore, a large “window current” might flow in the voltage range from −70 to −20 mV. In conclusion, in late post-myocardial infarction, a Na+current with specific kinetics and pharmacology may provide inward charges in a critical range of membrane voltages that are able to alter action potential time course and trigger ventricular arrhythmia. These apparent new characteristics of the Na+channel might result in part from environmental changes during heart remodeling.

The effects of chloride ions in excitation-contraction coupling and sarcoplasmic reticulum calcium release in twitch muscle fibre.

Using the sucrose vaseline gap technique, experiments were carried out on isolated frog twitch muscle fibre to investigate the role of chloride ions in excitation-contraction coupling. In current clamp conditions, replacement of chloride ions by impermeant anions led to an increase of the amplitude of the early after potential and of the amplitude of the twitch. Addition of a chloride channel blocker, anthracene-9-carboxylic acid gave similar results. In voltage clamp conditions, replacement of chloride ions by impermeant anions induced a decrease of the outward current and an increase of both the amplitude of the contraction and of the resting tension. Addition of anthracene-9-carboxylic acid gave similar results except that resting tension was not modified. Replacement of chloride ions by impermeant anions resulted in a shift of the tension-voltage relationship toward negative potentials and in an increase of the amplitude of the contraction at all potentials. Outward currents were also reduced at all potentials but no shift of the current-voltage relationship was observed. Similar results were obtained upon addition of anthracene-9-carboxylic acid. Rapid filtration experiments were performed on isolated sarcoplasmic reticulum vesicles to study the role of chloride ions in Ca2+ release. Under conditions where KCl was present in the intra- and extravesicular media, removal of chloride ions from the release solution produced a 2-fold increase in the rate of Ca(2+)-induced Ca2+ release. Together, these results suggest that, besides their involvement in the action potential time course, chloride ions could exert a negative control on the sarcoplasmic reticulum Ca2+ release.(ABSTRACT TRUNCATED AT 250 WORDS)

Time course of action potentials recorded from single human afferents.

It was the object of this study to measure the time course of the action potential in individual human sensory nerve fibres in relation to conduction properties of the axons. For this purpose, the technique of percutaneous microneurography was combined with intradermal electrical stimulation of distal portions of the axons. Recordings were made at the wrist level from 57 type-identified mechanoreceptive median nerve afferents [mainly rapidly adapting (RA) and slowly adapting type I (SAI)] innervating the glabrous skin of the hand. Measurements were made of the duration and time-to-peak of the positive peak of the diphasic (large positive phase followed by smaller, slower negative phase) action potential typically recorded using microneurography. Durations ranged from 0.31 to 0.75 ms (mean 0.50 ms) and times-to-peak from 0.12 to 0.45 ms (mean 0.21 ms), with no difference between afferent categories (RA, SAI). Time-to-peak was strongly positively correlated with duration (linear r = 0.81). Conduction velocity was measured over the distance extending from the point of intradermal stimulation (typically in the fingertips) to the point of recording at the wrist (distal conduction velocity). Absolute refractory period was measured using paired stimuli applied at the point of intradermal stimulation, within the receptive field of the afferent (distal absolute refractory period). Distal conduction velocities ranged from 15 to 60 m/s (mean 33 m/s), and distal refractory periods from 0.7 to 4.5 ms (mean 2.1 ms), with no difference between afferent types (RA, SAI). Distal absolute refractory period was inversely correlated with distal conduction velocity. The data were slightly better described assuming a non-linear (exponential) relationship; the non-linear correlation coefficient was -0.77. The time course of the action potential varied inversely with distal conduction velocity and directly with distal absolute refractory period. The time-to-peak versus conduction velocity data were slightly better described by a power than a linear relationship. Coefficients of correlation were: duration versus conduction velocity, linear r = -0.76; time-to-peak versus conduction velocity, non-linear r = -0.64; duration versus absolute refractory period, r = 0.70; time-to-peak versus absolute refractory period, r = 0.76. Extensive intercorrelation between the variables duration, distal conduction velocity and absolute refractory period was revealed by multiple correlation techniques. Inter- and intra-subject skin temperature variation was within 5 degrees C. Correcting the time course, conduction velocity and absolute refractory period values for temperature variation within this limited range did not affect the results.(ABSTRACT TRUNCATED AT 400 WORDS)

Electrophysiology of dopamine-denervated striatal neurons

Unilateral 6-hydroxydopamine-induced lesions of the substantia nigra have been used as an experimental model for Parkinson's disease. Although the biochemical and the behavioural effects of striatal denervation have been widely characterized, the physiological and pharmacological changes caused by dopamine depletion at the cellular level are still unknown. We studied the electrical activity of single rat striatal neurons recorded intracellularly in vitro from a brain slice preparation. Recordings were obtained at different periods after the denervation (4, 6, 8 months). In dopamine-denervated slices, unlike naive slices, most of the neurons showed spontaneous depolarizing postsynaptic potentials. The percentage of cells showing spontaneous depolarizing postsynaptic potentials was maximal 4 months after the denervation. In most of the dopamine-denervated neurons (60%) spontaneous depolarizing postsynaptic potentials were reversibly blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 microM), an antagonist of non-N-methyl-D-aspartate glutamate receptors. In some neurons, however, the amplitude of spontaneous depolarizing postsynaptic potentials was reduced by bicuculline (30 microM) suggesting that they were mediated by the release of endogenous gamma-aminobutyric acid (GABA). Intrinsic membrane properties (membrane potential, input resistance and firing pattern) and postsynaptic responses to different agonists of excitatory amino acid receptors were not altered in neurons recorded from dopamine-depleted slices. In dopamine-depleted slices, unlike in naive slices, LY 171555 (0.1-10 microM), a D2 dopamine receptor agonist, reduced the frequency and the amplitude of CNQX-sensitive spontaneous depolarizing postsynaptic potentials and reduced the amplitude of glutamate-mediated synaptic potentials evoked by cortical stimulation. LY 171555 did not affect the membrane responses to exogenous glutamate. SKF 38393 (3 microM), a D1 dopamine receptor agonist, decreased postsynaptic excitability of striatal neurons recorded from naive animals. On the contrary, this agonist was ineffective in most of the cells obtained from dopamine-depleted slices. These results suggest that dopamine-denervation augments neuronal excitability in the striatum. Abnormal excitability of striatal neurons is not caused by changes of the intrinsic membrane properties of these cells, but is the result of increased glutamatergic cortical inputs to the striatum. Dopamine-denervation also alters the physiological responses to dopamine receptor stimulation. Nigral lesions induce supersensitivity of D2 dopamine receptors controlling the release of glutamate and reduce the inhibitory influence of D1 receptors at postsynaptic level. These functional changes of the striatal neurons may alter the output signals from the striatum to the other structures of the basal ganglia and then produce most of the physiopathological changes observed in Parkinson's disease.

Method and theory of monophasic action potential recording

MAP recordings have been at the cradle of cardiac electrophysiology but only recently, through safer and simpler technology, have gained wider access to clinical electrophysiology. In contrast to conventional electrode catheter recordings, MAP recording devices provide precise information not only of the local activation time but of the entire local repolarization time course as well. Although the MAP does not reflect the absolute amplitude or upstroke velocity of transmembrane action potentials, it delivers highly accurate information on the action potential duration and configuration, including early afterdepolarizations as well as relative changes in transmembrane diastolic and systolic potential changes. Based on available data, the MAP probably reflects the transmembrane voltage of cells within a few millimeters of the exploring electrode. MAPs can be recorded by catheter technique from the endocardial surface and by special probes from the epicardium in the operating room. The contact electrode technique is preferable over suction electrodes because it is safer and simpler to use in patients and because it produces more stable, longer-lasting signals. A modified contact MAP catheter incorporates pacing electrodes and permits simultaneous assessment of action potential duration and refractoriness. This not only facilitates the use of MAP catheters in routine electrophysiological studies but also is important for assessing the voltage-independent effects of antiarrhythmic drugs on refractoriness. MAP recordings offer the opportunity to study, in the in situ heart, a variety of pertinent electrophysiological phenomena including, for example, effects of cycle length changes and antiarrhythmic drugs on action potential duration or the role of afterdepolarizations in the genesis of triggered arrhythmias. Due to vigorous heart beating, movement artifacts may occur and need to be distinguished from true abnormalities in the action potential time course. With these limitations in mind, MAP recordings are a valuable addition to clinical electrophysiological studies.

GABA B agonists modulate a transient potassium current in cultured mammalian hippocampal neurons

Depolarization of voltage-clamped cultured rat hippocampal neurons from holding potentials more negative than −60 mV produced a transient outward current with the characteristics of an A-current: it was 50% inactivated at a holding potential of −85 mV and blocked by 4-aminopyridine (1 mM). In the presence of GABA or baclofen (50–200 μM), with or without bicuculline, inactivation of this current was shifted to more positive potentials so that there was little inactivation at −70 mV. Activation of the A-current was also shifted to more positive potentials by these agonists, but the voltage dependence of activation of the sodium current was unaffected. If A-currents with similar properties can influence the time course of action potentials in presynaptic terminals, GABA B agonists could make action potentials briefer by potentiating the A-current and hence depress transmitter release.

Paralysis of skeletal muscle by butanedione monoxime, a chemical phosphatase.

The chemical phosphatase butanedione monoxime (BDM) reversibly inhibited twitches and tetanic contractions in bundles of rat soleus fibres in a dose-dependent manner (2-20 mM) but had no effect on the amplitude or time course of action potentials. In addition, BDM reversibly reduced the amplitude of potassium contractures demonstrating a depressant effect on contraction not mediated by action potentials. BDM had no effect on asymmetric charge movement but depressed calcium currents across the surface membrane in voltage-clamped fibres. The most significant effect of BDM on excitation-contraction coupling was a reduction in the amplitude of the calcium transient associated with contraction in aequorin-injected fibres. While these experiments do not eliminate the possibility of a direct effect of BDM on contractile filaments, reduction of calcium release from the sarcoplasmic reticulum, at least at low concentrations of BDM (below 2 mM), would seem to be the main mechanism for the inhibition of contractions in rat skeletal muscle.

Chronic exposure to lead causes persistent alterations in the electric membrane properties of neurons in cell culture.

The effects of chronic lead (Pb) exposure on neuronal electric membrane properties (EMP) were determined using neural cell cultures of adult mouse dorsal root ganglia (DRG). Cultures were exposed to Pb concentrations ranging from 0 to 100 microM for 12 days (8 DIV to 20 DIV). EMP were determined in Pb-free medium either immediately after withdrawal (IWD), or 6 days after withdrawal (6WD) from Pb. For IWD, regression analysis indicated that a number of EMP varied significantly with increasing Pb concentration. The largest such change occurred for electrical excitability which decreased significantly with increasing Pb (P = 0.000), being reduced by approximately two-thirds for neurons exposed to 100 microM Pb; resting membrane potential increased with Pb (P = 0.000); membrane time constant decreased with Pb (P = 0.007); action potential afterhyperpolarization decreased with Pb (P = 0.023). There was also evidence that the time course of action potentials was accelerated with increasing Pb concentrations, the rate of fall of neurons with biphasic falling phases being particularly increased (P = 0.047). This general pattern of altered EMP was observed for the 6WD condition also, indicating that chronic exposure to Pb caused persistent abnormalities in neuronal membranes even after 6 days of cultivation in Pb-free medium. The patterns of alterations in EMP suggested that chronic Pb exposure caused a prolonged increase in potassium permeability. It was proposed that the latter was mediated through a Pb-induced increase in intracellular ionic calcium and the associated disruption of calcium homeostasis.

Rate-limiting step of inhibitory post-synaptic current decay in Aplysia buccal ganglia.

1. In neurones BL and BR 3, 6, 8, 9, 10 and 11 of Aplysia buccal ganglia, cholinergic inhibitory post-synaptic potentials are produced by activity in either of two presynaptic cells. In order to analyse the synaptic conductance change, neurones were voltage-clamped inhibitory post-synaptic currents (i.p.s.c.) recorded. 2. The synaptic conductance change rises to an average peak value of 0.65 micromho and decays exponentially with single time constant tau of 19 msec. 3. We have attempted to identify the rate-limiting step responsible for i.p.s.c. decay from among the following possibilities: (1) acetylcholine (ACh) supply, (2) ACh removal by diffusion, (3) ACh removal by hydrolysis or (4) a slow unbinding or conformational change closing open synaptic current channels. 4. Cooling prolongs tau, with Q10 of 5.2. Cooling and eserine treatment together produce greatly prolonged, exponentially decaying i.p.s.c.s with tau > 150 msec. These results suggest that ACh removal, either by diffusion or hydrolysis, is not the rate-limiting step. 5. Prolonging synaptic action potential time course with intracellular injection of tetraethylammonium broadens the i.p.s.c. peak but does not affect the decay tail, suggesting that the rate-limiting step is not ACh release. 6. The spectrum of ACh-induced current fluctuations is fitted by a double Lorentzian with cut-off frequencies of 7.8 and 47 Hz. The frequency of the slower component corresponds to the macroscopic i.p.s.c. decay tau. 7. We conclude that a slow conformational change closing open synaptic current channels is likely to determine i.p.s.c. decay. We cannot, however, exclude either delayed diffusion or a late tail of slow ACh release as possibilities.

The electrophysiological effects of disopyramide phosphate on canine ventricular muscle and Purkinje fibers in normal and low potassium.

We studied the effect of lowering the extracellular potassium concentration ([K+]o) on the electrophysiological actions of disopyramide phosphate, a new antiarrhythmic drug. At low [K+]o, therapeutic concentrations of disopyramide phosphate caused significantly less depression of action potential amplitude and maximum upstroke velocity of both Purkinje fiber and ventricular muscle action potentials. The drug shifted the membrane responsiveness curve along the voltage axis to more negative membrane potentials regardless of [K+]o. However, a greater shift occurred when [K+]o was normal. Disopyramide phosphate prolonged both action potential duration and effective refractory period in all fibers but there was consistently greater prolongation of these parameters at low [K+]o. More importantly, disopyramide phosphate altered repolarization time course of action potentials in such a way that action potentials with dissimilar durations throughout the ventricular conducting system became more equal. The drug was less effective in decreasing this disparity in action potential durations throughout the ventricles in the presence of low [K+]o. These modifications of the electrophysiological actions of disopyramide by low [K+]o suggest that a therapeutic concentration of disopyramide might have less of an antiarrhythmic effect in the presence of hypokalemia.

Effects of pH on excitation and contraction in frog twitch muscle.

The electrical and mechanical behaviour of frog twitch muscle in response to changes in membrane potential has been examined over a wide range of hydrogen ion concentration (pH 3.0–11.0). The changes in resting and action potentials, twitches, and maximum potassium-induced contractures (K contractures) were remarkably small when the pH was varied between 5.0 and 10.0. The time course of action potentials generally displayed small graded changes with variation in pH, possibly as the result of changes in surface potential.The amplitude of twitches and maximum K contractures was substantially decreased when pH was reduced to 4.0 or raised to 11.0 without significant alteration in membrane resting potential or consistent suppression of excitation, but maximum caffeine-induced contractures were unchanged. Replacement of chloride with perchlorate promptly antagonized the depressant effects of pH extremes (4.0, 11.0) on both twitch amplitude and maximum K-contracture tension. Acid-induced reductions in maximum K-contracture tension also were partially antagonized by increased calcium concentration. The onset and recovery from the contraction-depressant effects of pH extremes were too slow to be explained by the titration of groups immediately accessible at the membrane surface but too rapid to be accounted for by changes in intracellular pH. Thus, excitation and contraction apparently were uncoupled by sufficient alteration in extracellular pH. Changes in external pH had little effect on the impairment of maximum K contractures by media lacking divalent cations, or on the restoration of such responses by perchlorate except at very alkaline pH (10.0–11.0).The threshold for K contractures was reduced at pH 11.0, but otherwise was little affected by variation in pH at normal concentrations of divalent cations. Altered pH did not modify the usual effects of increased calcium concentration on the relation between potassium concentration and K-contracture tension. When K contractures were maintained by perchlorate in the absence of divalent cations, hydrogen ions displayed calcium-like actions on the relation between external K concentration ([K]0) and K-contracture tension, and also on the time course of submaximum K contractures. These observations are compatible with similar effects of hydrogen and calcium ions on surface potential.The problem of identifying putative charged groups which might influence the linkage between contractile responses and changes in membrane potential is discussed.